Bispecific hiv-1-neutralizing antibodies

ABSTRACT

In various embodiments, the present invention relates generally to using bispecific antibodies in the prevention and treatment of HIV.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation-in-part of U.S. patent applicationSer. No. 15/414,822, filed Jan. 25, 2017, which is a divisional of U.S.patent application Ser. No. 14/558,341, filed Dec. 2, 2014, which ispatented as U.S. Pat. No. 9,587,012, which claims priority to and thebenefit of U.S. Provisional Patent Application No. 61/910,685, filedDec. 2, 2013, each of which is hereby incorporated by reference in itsentirety.

FIELD OF THE INVENTION

In various embodiments, the present invention relates generally to usingbispecific antibodies in the prevention and treatment of HIV.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted in ASCII format via EFS-Web and is hereby incorporated byreference in its entirety. Said ASCII copy, created Dec. 21, 2017, isnamed ADR-001CP_ST25.txt and is 145,851 bytes in size.

BACKGROUND

Passive immunization with antibodies (Abs) is a recognized method ofprophylaxis and treatment of infectious diseases. This approach mayinvolve preparing human immunoglobulins from donors who recovered froman infectious disease and utilizing such preparations, containing Absspecific for the infectious organism, to protect a recipient against thesame disease. Alternatively, therapeutic antibodies can be made byimmunizing mice with an antigen, and then engineering/humanizing themouse Ab into a human version. Monoclonal antibodies (mAbs) arehomogeneous in terms of physical characteristics and immunochemicalreactivity, and so offer the possibility of absolute specific activity.

That specificity can ultimately be a limitation for some targets, sopractitioners have developed “bispecific” mAbs composed of fragments oftwo different mAbs and which bind to two different types of antigen.This facilitates binding to antigens expressed only weakly, for example.Some bispecific mAbs can stimulate strong immune responses, limitingtheir clinical application. One recent approach to ameliorating thiseffect is “CrossMab” methodology, a bispecific antibody format thatadopts a more native antibody-like structure.

The prospects for generating a highly potent bispecific or bivalentantibody against a pathogen, such as HIV, for clinical use involves manyuncertainties. The low spike density and spike structure on HIV mayimpede bivalent binding of antibodies to HIV, for example, and thegeometry and spatial relationship of cell surface anchoring are notwell-characterized. Nor is it known whether sufficient epitopeaccessibility on the HIV envelope exists. CrossMab bispecific antibodiesthat are anchored to a host cell membrane offer the possibility ofimproved local antibody concentration, targeting of sequential and/orinterdependent entry steps, and compensating for monovalent binding.

Further still, large-scale, commercial production of antibodies remainschallenging. For example, the production of therapeutic antibodies oftenrequires the use of very large cell cultures followed by extensivepurification steps, under Good Manufacturing Practice conditions,thereby resulting in extremely high production costs. Other limitationssuch as poor insolubility, protein aggregation, and protein instabilitycan also make manufacturing of antibodies less than optimal.

Accordingly, there remains a need for therapeutically effective HIVantibodies that can be easily produced at a commercial scale.

SUMMARY

In one aspect, the present invention pertains to a bispecific antibodyfor neutralizing HIV. The bispecific antibody includes portions of afirst and a second antibody, in which the first antibody binds to a HIVenvelope protein. In certain embodiments, the first antibody is selectedfrom PGT145, PG9, PGT128, PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10,10E8 and a variant thereof. In certain embodiments, the bispecificantibody includes portions of a second antibody, in which the secondantibody binds to a cell membrane protein. For example, the secondantibody binds to a cell receptor protein or a cell membrane co-receptorprotein. In an embodiment, the second antibody is selected from a CD4antibody, a CCR5 antibody and a CXCR4 antibody, such as Pro 140,ibalizumab, 515H7, or a variant thereof. In various embodiments, thebispecific antibody has a CrossMab format.

In another aspect, the present invention provides a bispecific antibodyincluding portions of a first antibody and a second antibody, whereinthe first antibody binds to a HIV envelope protein and the secondantibody binds to a cell membrane protein. In various embodiments, thebispecific antibody has a CrossMab format.

In various embodiments, pharmaceutical compositions including thebispecific antibodies disclosed herein are also provided. Thepharmaceutical composition may be formulated for oral, intranasal,pulmonary, intradermal, transdermal, subcutaneous, intramuscular,intraperitoneal, or intravenous delivery.

In a further aspect, methods for neutralizing HIV are provided. Themethods include the steps of contacting an antigen binding site with abispecific antibody that binds a HIV envelope protein and contactinganother antigen binding site with a bispecific antibody that binds acell membrane protein.

In another aspect, methods for treating a patient infected with HIV arealso provided. The methods include administering to the patient any ofthe bispecific antibodies or pharmaceutical compositions as disclosedherein. In an embodiment, the patient is human.

BRIEF DESCRIPTION OF THE DRAWINGS

In the drawings, like reference characters generally refer to the sameparts throughout the different views. Also, the drawings are notnecessarily to scale, with an emphasis instead generally being placedupon illustrating the principles of the invention. In the followingdescription, various embodiments of the present invention are describedwith reference to the following drawings, in which:

FIG. 1 is a diagram illustrating a CrossMab antibody derived from twoIgG monoclonal antibodies.

FIG. 2A is a diagram illustrating an iMab antibody (shorthand for themonoclonal antibody ibalizumab) that targets CD4 and a Pro 140 antibodythat targets CCR5.

FIG. 2B is a diagram illustrating mAbs that target the HIV envelopegp120.

FIG. 3 is a graph comparing the maximum percentage inhibition (MPI)against cell-to-cell HIV transmission using a combination of iMab and10E8 antibodies with CrossMab bispecific 10E8/iMab antibodies. Exceptotherwise stated, all iMab-based bispecific antibodies were constructedusing the MV1 variant.

FIGS. 4A-J are a series of graphs comparing the inhibition of variousstrains of X4 and dual-tropic HIV using varying concentrations of 10E8,Pro 140 or 10E8/P140 antibodies. P140 is shorthand for Pro 140.

FIGS. 5A-G are a series of graphs comparing the inhibition of variousstrains of HIV using varying concentrations of 10E8, Pro 140, 10E8/P140or a combination of the individual 10E8 and Pro 140 monoclonalantibodies.

FIGS. 6A-D are a series of graphs comparing the inhibition of variousstrains of HIV using varying concentrations of 10E8, X19, 10E8/X19 or10E8/P140 antibodies.

FIGS. 7A-H are a series of graphs comparing the inhibition of variousstrains of HIV using varying concentrations of 10E8, Pro 140, 10E8/P140and 10E8/αHer2 antibodies.

FIG. 8A is a graph comparing the binding of CrossMab bispecificantibodies 10E8/iMab and Δ10E8/iMab to the HIV-1 glycoprotein MPER.

FIGS. 8B-E are a series of graphs comparing the inhibition percentagesof 10E8 (light gray lines) and Δ10E8 (dark gray lines) against iMabresistant R5 viruses (FIG. 8B) and X4 viruses (FIG. 8C), as well as theinhibition percentages of 10E8/iMab (light gray lines) and Δ10E8/iMab(dark gray lines) against iMab resistant R5 viruses (FIG. 8D) and X4viruses (FIG. 8E).

FIGS. 9A-G are a series of graphs comparing the inhibition of variousstrains of HIV using varying concentrations of 10E8, Δ10E8, 4E10,10E8/P140, Δ10E8/P140 and 4E10/P140 antibodies.

FIG. 10 is a graph comparing the antiviral coverage of the CrossMabantibodies 10E8/Pro140 and 10E8/iMab, their parental monoclonalantibodies 10E8, Pro140 and iMab, and various other HIVenvelope-targeting monoclonal antibodies against a large panel of HIVenvelope pseudotyped viruses.

FIGS. 11A-E are a series of graphs comparing the maximum percentageinhibition (MPI) of a large panel of HIV envelope pseudotyped viruseswith the monoclonal antibody iMab (grey bars in all panels) and theCrossMab antibodies PGT145/ibalizumab (145/iMab; FIG. 11A),PGT128/ibalizumab (128/iMab; FIG. 11B), PGT151/ibalizumab (151/iMab;FIG. 11C), 3BNC117/ibalizumab (117/iMab; FIG. 11D) and 10E8/ibalizumab(10E8/iMab; FIG. 11E).

FIGS. 12A-E are a series of graphs comparing the maximum percentageinhibition (MPI) and IC80 antibody concentrations of the CrossMabantibodies PGT145/ibalizumab (145/iMab; FIG. 12A), PGT128/ibalizumab(128/iMab; FIG. 12B), PGT151/ibalizumab (151/iMab; FIG. 12C),3BNC117/ibalizumab (117/iMab; FIG. 12D) and 10E8/ibalizumab (10E8/iMab;FIG. 12E) against a large panel of HIV envelope pseudotyped viruses.

FIGS. 13A-E are a series of graphs comparing the IC80 antibodyconcentrations for iMab- and Pro140-based CrossMab bispecific antibodiesand their parent antibodies for PGT145/iMab and PGT145/Pro140 (FIG.13A), 3BNC117/iMab and 3BNC117/Pro140 (FIG. 13B), PGT128/iMab andPGT128/Pro140 (FIG. 13C), PGT151/iMab and PGT151/Pro140 (FIG. 13D) and10E8/iMab and 10E8/Pro140 (FIG. 13E).

FIGS. 14A-E are a series of graphs comparing the IC50 antibodyconcentrations for iMab- and Pro140-based CrossMab bispecific antibodiesand their parent antibodies for PGT145/iMab and PGT145/Pro140 (FIG.14A), 3BNC117/iMab and 3BNC117/Pro140 (FIG. 14B), PGT128/iMab andPGT128/Pro140 (FIG. 14C), PGT151/iMab and PGT151/Pro140 (FIG. 14D) and10E8/iMab and 10E8/Pro140 (FIG. 14E).

FIGS. 15A-E are graphs displaying the IC80 antibody concentrations foriMab-based CrossMab bispecific antibodies and their parent antibodiesagainst cell-to-cell transmission of HIV for 10E8/iMab (FIG. 15A),3BNC117/iMab (FIG. 15B), PGT145/iMab (FIG. 15C), PGT128/iMab (FIG. 15D)and PGT151/iMab (FIG. 15E).

FIG. 16 is a graph displaying the maximum percent inhibition (MPI) ofCrossMab bispecific antibodies and parental antibodies againstcell-to-cell transmission of HIV.

FIG. 17A is a graph comparing the inhibition of an HIV strain againstvarying concentrations of 10E8, Pro 140, 10E8/P140 CrossMab bispecificantibody, and a combination of individual 10E8 and Pro 140 monoclonalantibodies.

FIG. 17B is a graph comparing the inhibition of an HIV strain againstvarying concentrations of iMab, 10E8, 10E8/iMab CrossMab bispecificantibody, and a combination of individual 10E8 and iMab monoclonalantibodies.

FIGS. 18A-D are a series of graphs comparing the inhibition of variousHIV R5 strains against varying concentrations of 10E8, Pro140, 10E8/P140and 10E8/515H7 antibodies.

FIGS. 18E-H are a series of graphs comparing the inhibition of variousHIV X4 strains against various concentrations of 10E8, 515H7 and10E8/515H7 antibodies.

FIGS. 19A-B are a series of graphs comparing inhibition of various HIVstrains against varying concentrations of 10E8/Pro140, 10E8/iMab,10E8/515H7 and 10E8/X19 antibodies.

FIG. 19C indicates the density of CD4, CCR5 and CXCR4 receptors presenton TZM-bl cells.

FIG. 20 compares the binding of CrossMab bispecific antibodies10E8/Pro140, Δ10E8/Pro140 and 4E10/Pro140 to the HIV-1 glycoproteinMPER.

FIGS. 21A-G are a series of graphs comparing the inhibition of variousstrains of HIV against varying concentrations of 4E10, Pro140 and4E10/P140 and 10E8/P140 antibodies.

FIG. 22A is size exclusion chromatography analysis of the CrossMabantibodies 10E8/iMab, 10E8/P140 and 3BNC117/iMab.

FIG. 22B is size exclusion chromatography analysis of the monoclonalantibodies iMab, 10E8 and Pro140.

FIG. 23 is size exclusion chromatography analysis of monoclonal antibody10E8 and a chimeric antibody comprised of the 10E8 heavy chain pairedwith the 4E10 light chain.

FIGS. 24A-C are a series of size exclusion chromatography graphs of: themonoclonal antibodies 10E8 and 4E10 and a chimeric antibody comprised ofthe 10E8 heavy chain paired with the 4E10 light chain (FIG. 24A), themonoclonal antibody 10E8 and 10E8 mutants with potentially stabilizingmutations genetically engineered in the 10E8 light chain (FIG. 24B), andthe monoclonal antibody 10E8 and 10E8 mutants genetically grafted withthe kappa light chain of non-10E8 antibodies (FIG. 24C).

FIG. 25 is a size exclusion chromatography graph of the monoclonalantibody 4E10 and 4E10 mutants genetically grafted with the lightregions of 10E8 that included the CDR1 region, CDR2 region, CDR3 region,or combined CDR1, CDR2 and CDR3 regions.

FIG. 26A is a size exclusion chromatography graph of 10E8 chimericantibodies. CDR123 is a chimeric antibody of the 10E8 heavy chain pairedwith a 10E8 light chain genetically grafted with the 10E8 antibodygermline CDR region sequences. FW123 is a chimeric antibody of the 10E8heavy chain paired with a 10E8 light chain genetically grafted with the10E8 antibody germline framework region sequences.

FIG. 26B is a table indicating the expression, HIV MPER binding ability,size exclusion chromatography profile, and HIV neutralization profile ofthe CDR123 and FW123 antibodies.

FIG. 27 is a size exclusion chromatography graph of monoclonal antibody10E8, its somatic variant H6L10, and a CrossMab bispecific antibodyconsisting of H6L10 paired with Pro140.

FIG. 28 is a graph depicting the pharmacokinetics profiles of 10E8,H6L10/Pro 140 and its parental antibodies in a mouse model.

FIG. 29 is a graph comparing the potency of 10E8_(v 1.0)/iMab or P140CrossMab antibodies with 10E8/iMab or P140 antibodies.

FIG. 30 is a graph depicting the pharmacokinetics of 10E8 and CrossMabantibodies derived from several 10E8 variants and iMab or P140 in amouse model.

FIGS. 31A-B are a series of graphs depicting the HIV viral coverage of10E8_(v1.1)/P140 and 10E8_(v2.0)/iMab antibodies.

FIGS. 31C-D are a series of graphs depicting size exclusionchromatography stability graphs of 10E8_(v1.1)/P140 and 10E8_(v2.0)/iMabantibodies.

FIGS. 32A-B are a series of graphs depicting the size exclusionstability graphs of 10E8_(v1.1)/P140 and 10E8_(v2.0)/iMab antibodiesstored in PBS at 4° C.

FIG. 33 depicts a native mass spectroscopy analysis of the10E8_(v2.0)/iMab (N297A) antibody.

FIGS. 34A-C are a series of graphs comparing the activity of10E8_(v1.1)/P140 and 10E8_(v2.0)/iMab on a HIV Clade C panel, and theIC50 and IC80 activities of the antibodies.

FIGS. 35 and 36 are graphs comparing the potency of 10E8_(v1.1)/P140,10E8_(v2.0)/iMab, and various monoclonal antibodies against HIV.

FIGS. 37A-C demonstrate that a select number of 10E8V2.0/iMab (alsoreferred to as 10E8.2/iMab) variants retained functional antiviralactivity and increased solubility. FIG. 37A demonstrates that some ofthe 10E8.2/iMab variants retained functional activity in an in vitroHIV-1 neutralization assay. FIG. 37B shows that 10E8.2/iMab and some ofthe 10E8.2/iMab variants have similar in vivo pharmacokinetic profiles.FIG. 37C shows the precipitation profiles of 10E8.2/iMab and some of the10E8.2/iMab variants under thermal stress-inducing conditions.

FIGS. 38A-B show the results of size exclusion chromatography, which wasused to identify 10E8.2/iMab variants with the least aggregation afterthermal stress-inducing conditions.

FIGS. 39A-B illustrate the solubility of 10E8.2/iMab and the 10E8.4/iMabvariant at 4° C. after ultracentrifugation.

FIG. 40 demonstrates the turbidity of 10E8.2/iMab and the 10E8.4/iMabvariant at different concentrations over time.

FIG. 41 shows the thermostability of 10E8.2/iMab and the 10E8.4/iMabvariant as assessed by differential scanning calorimetry.

FIG. 42 demonstrates the turbidity of 10E8.2/iMab and the 10E8.4/iMabvariant after forced degradation at 50° C. for six days. For each set ofhistograms, the bars from left to right represent 10E8.2/iMab(pre-centrifugation), 10E8.4/iMab (pre-centrifugation), 10E8.2/iMab(post-centrifugation), and 10E8.4/iMab (post-centrifugation).

FIG. 43 shows the anti-HIV coverage of 10E8.2/iMab and the 10E8.4/iMabvariant.

FIG. 44 is a graph showing the in vivo antiviral activity of 10E8.2/iMaband the 10E8.4/iMab variant in a humanized mouse model of HIV-1infection.

FIG. 45A, shows a sequence alignment of the light chains of 10E8.2/iMab(SEQ ID NO:33) and the 10E8.4/iMab variant (SEQ ID NO:44).

FIG. 45B shows a sequence alignment of the heavy chains of 10E8.2/iMab(SEQ ID NO:34) and the 10E8.4/iMab variant (SEQ ID NO:42). Underlinedsequences denote CDR1, CDR2, and CDR3. Italicized sequences denoteconstant light chain or constant heavy chain sequences.

FIG. 46 is a graph showing exemplary variants of 10E8 antibodies thatare stable while retaining anti-HIV activity.

DETAILED DESCRIPTION

Embodiments of the present invention provide for inhibition of HIV. Invarious implementations, bispecific antibodies are formed, eachincluding heavy chain and light chain components from two differentparent antibodies. In various embodiments, one parent antibodyspecifically binds HIV, for example, the HIV envelope protein Env. Invarious embodiments, the other parent antibody specifically binds a cellmembrane protein, for example CD4 and CCR5.

In various embodiments, the bispecific antibody (e.g., a HIV CrossMabantibody) of the present invention has the natural architecture of anIgG molecule, but with bispecificity. In a bispecific antibody, a heavychain and light chain from each of two parental antibodies are combined,providing an antibody in which the antigen binding sites of fragmentantigen-binding 1 (Fab1) and Fab2 have different binding specificities.In certain embodiments, the bispecific antibody is a CrossMab formatantibody, as shown in FIG. 1. In a CrossMab format, one heavy chainincludes a “knob” structure and the other heavy chain includes acorresponding “hole” structure, and the positions of the constantdomains (i.e., CL and CH1) from one parental antibody are switched,which together ensure correct pairing of heavy chains and light chainsduring assembly.

Various mAbs have been shown to block HIV infection by targeting andbinding to the HIV envelope protein Env (FIGS. 2B and 10). These mAbsinclude, for example, PGT145, PG9, PGT128, PGT121, 10-1074, 3BNC117,VRC01, PGT151, 4E10, and 10E8. FIG. 2B (adapted fromwww.scripps.edu/news/press/2014/20140424hiv.html) illustrates how themAb PGT145 targets the V1/V2 epitope on the HIV viral envelope gp120;how mAb PGT128 targets the glycan on the V3 stem region of HIV gp120;how mAb 3BNC117 targets the CD4 binding site of HIV gp120; how mAb 10E8targets the membrane proximal external region (MPER) of HIV gp41; andhow mAb PGT151 targets an epitope on both HIV gp120 and HIV gp41.

In addition, monoclonal antibodies Pro 140 (“P140”), Ibalizumab (“iMab”)and 515H7 have been shown to block HIV infection by targeting andbinding to CCR5, CD4 and CXCR4 human cell membrane proteins,respectively (FIG. 2A). Specifically, FIG. 2A shows how iMab targetsCD4, the primary receptor for HIV-1 entry that is expressed on humanT-cells; and how Pro 140 targets CCR5, a co-receptor for HIV-1 entry byCCR5 tropic HIV-1.

Although the ensuing discussion focuses on the use of bispecificantibodies directed to Env and the cell membrane proteins CD4 and CCR5,it is to be understood that this is solely for ease of presentation, andthat any suitable antibody directed to any HIV epitope and any suitableantibody directed to any suitable cell membrane protein may be used andare within the scope of the invention.

Accordingly, in various embodiments, the present invention providesbispecific antibodies that target and bind to the HIV Env protein aswell as the cell membrane proteins CCR5, CD4 and/or CXCR4. In certainembodiments, the bispecific antibodies include sequences (for example,heavy and light chain sequences) derived from, but not limited to, thePGT145, PG9, PGT128, PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10,and/or 10E8 antibodies and variants thereof.

The amino acid sequences defining the heavy and light chains of thePGT145 antibody can be found, for example, atwww.ncbi.nlm.nih.gov/protein/3U1S_H andhttp://www.ncbi.nlm.nih.gov/protein/3U1S_L, respectively, the entirecontents of which are incorporated herein by reference.

The amino acid sequences defining the heavy and light chains of the PG9antibody can be found, for example, at www dot ncbi dot nlm dot nih dotgov/protein/3U4E_H and www dot ncbi dot nlm dot nih dotgov/protein/3MUH_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of thePGT128 antibody can be found, for example, at www dot ncbi dot nlm dotnih dot gov/protein/3TYG_H and www dot ncbi dot nlm dot nih dot govprotein/3TYG_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of thePGT121 antibody can be found, for example, at www dot ncbi dot nlm dotnih dot gov/protein/4FQC_H and www dot ncbi dot nlm dot nih dotgov/protein/4FQC_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of the10-1074 antibody can be found, for example, in Mouquet H., et al.,(2012) PNAS, 109(47): E3268-77 (including supplementary information),the entire contents of which are incorporated herein by reference.

The amino acid sequences defining the heavy and light chains of the3BNC117 antibody can be found, for example, at www dot ncbi dot nlm dotnih dot gov/protein/4LSV_H and www dot ncbi dot nlm dot nih dotgov/protein/4LSV_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of theVRC01 antibody can be found, for example, at www dot ncbi dot nlm dotnih dot gov/protein/4LST_H and www dot ncbi dot nlm dot nih dotgov/protein/4LST_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of thePGT151 antibody can be found, for example, at www dot ncbi dot nlm dotnih dot gov/protein/4NUG_H and www dot ncbi dot nlm dot nih dotgov/protein/4NUG_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of the 4E10antibody can be found, for example, at www dot ncbi dot nlm dot nih dotgov/protein/4LLV_H and www dot ncbi dot nlm dot nih dotgov/protein/4LLV_L, respectively, the entire contents of which areincorporated herein by reference.

The amino acid sequences defining the heavy and light chains of the 10E8antibody can be found, for example, at www dot ncbi dot nlm dot nih dotgov/protein/4G6F_B and www dot ncbi dot nlm dot nih dotgov/protein/4G6F_D, respectively, the entire contents of which areincorporated herein by reference.

In certain embodiments, the bispecific antibodies include sequences (forexample, heavy and light chain sequences) derived from, but not limitedto, the P140, iMab (or the MV1 variant) and/or 515H7 antibodies andvariants thereof. The heavy and light chain sequences of the Pro 140,iMab (or its MV1 variant), and 515H7 antibodies are further described,for example, in Olson, W. C. et al., (1999) J Virol., 73(5):4145-55,Trkola, A. et al., (2001) J Virol., 75(2):579-88, U.S. Pat. No.7,122,185, Burkly L. C. et al., (1992) J Immunol., 149(5):1779-87, MooreJ. P. et al., (1992) J Virol., 66(8):4784-93, Reimann K. A., et al.,(1997) AIDS Res Hum Retroviruses, 13(11):933-43, International PatentPublication No. WO2014100139, and European Patent Publication No.EP2246364, the entire contents of all of which are incorporated hereinby reference.

As used herein, an antibody “variant” refers to an antibody which has anamino acid sequence which differs from the amino acid sequence of aparent antibody from which it is derived. In various embodiments, thevariant has one or more amino acid alterations with respect to theparent antibody.

In various embodiments, the bispecific antibody of the present inventionincludes a heavy and light chain sequence from the PGT145, PG9, PGT128,PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10, or 10E8 antibody or avariant thereof and a heavy and light chain sequence from the P140, iMab(or the MV1 variant), or 515H7 antibody or a variant thereof.

In exemplary embodiments, a series of HIV CrossMab antibodies have beenconstructed including but not limited to, for example, 145/MV1, 117/MV1,128/MV1, 10E8/MV1, 145/P140, 128/P140, 117/P140, 10E8/P140,10E8/alpha-Her2, 10E8/X19, and 4E10/P140. PGT145 (“145”), 3BNC117(“117”), PGT128 (“128”), and 10E8 are four different HIV envelopeantibodies. Pro 140 (“P140”) is a mAb that binds to the cell surfacereceptor CCR5. MV1 is a CD4 antibody that is a modified variant of themAb Ibalizumab (“iMab”; see, for example, International PatentPublication No. WO2014100139, incorporated herein by reference in itsentirety). X19 is one of the antibody variants of the anti-cell surfacereceptor CXCR4 (see, for example, U.S. Pat. No. 8,329,178, incorporatedherein by reference in its entirety) that does not bind to cellsexpressing CXCR4 (and is therefore used as a non-surface bindingcontrol). Alpha-Her2 is a mAb that binds to the Her2 receptor expressedon cells. Many of these CrossMab antibodies increase the breadth of HIVneutralization as compared to their parental antibodies (i.e.,monoclonal antibodies MV1, 145, 117 or 10E8). In various embodiments,the bispecific antibodies of the invention significantly increase thepotency of neutralization against HIV as compared to their parentalantibodies.

The amino acid sequences defining the heavy and light chains of variousexemplary HIV CrossMab antibodies are shown below.

145/MV1 Antibody:

Amino Acid Sequence Defining the MV1 Derived Light Chain of the 145/MV1Antibody—MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the 145/MV1Antibody—MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the PGT145 Derived Light Chain of the145/MV1 Antibody—PGT145-LC (SEQ ID NO:3):

EVVITQSPLFLPVTPGEAASLSCKCSHSLQHSTGA NYLATNYLQRPGQTPRLLIHLATHRASGVPDRFSGSGSGTDFTLKISRVESDDVGTYYCMQGLHSPTNTF GQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQTNKVDNALQSGNSQESVTEQD SKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

Amino Acid Sequence Defining the PGT145 Derived Heavy Chain of the145/MV1 Antibody—PGT145-HC-Knob (SEQ ID NO:4):

QVQLVQSGAEVKKPGSSVKVSCKASGNSFSNHDVH WVRQATGQGLEWMGWMSHEGDKTGLAQKFQGRVTITRDSGASTVYMELRGLTADDTAIYYCLTGSKHRLR DYFLYNEYGPNYEEWGDYLATLDVWGHGTAVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPE PVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPR EEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQ VSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEAL HSHYTQKSLSLSPGK

117/MV1 Antibody:

Amino Acid Sequence Defining the MV1 Derived Light Chain of the 117/MV1Antibody—MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the 117/MV1Antibody—MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the 3BNC117 Derived Light Chain of the117/MV1 Antibody—3BNC117-LC (SEQ ID NO:5):

DIQMTQSPSSLSASVGDTVTITCQANGYLNWYQQR RGKAPKLLIYDGSKLERGVPSRFSGRRWGQEYNLTINNLQPEDIATYFCQVYEFVVFGQGTKVQVDIKRT VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

Amino Acid Sequence Defining the 3BNC117 Derived Heavy Chain of the117/MV1 Antibody—3BNC117-HC-Knob (SEQ ID NO:6):

QVQLLQSGAAVTKPGASVRVSCEASGYNIRDYFIH WWRQAPGQGLQWVGWINPKTGQPNNPRQFQGRVSLTRHASWDFDTFSFYMDLKALRSDDTAVYFCARQRS DYWDFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHT FPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPS VFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVL HQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPR

128/MV1 Antibody:

Amino Acid Sequence Defining the MV1 Derived Light Chain of the 128/MV1Antibody—MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the 128/MV1Antibody—MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the PGT128 Derived Light Chain of the128/MV1 Antibody—PGT128-LC (SEQ ID NO:7):

QSALTQPPSASGSPGQSITISCIGTSNNFVSWYQQ HAGKAPKLVIYDVNKRPSGVPDRFSGSKSGNTASLTVSGLQTDDEAVYYCGSLVGNWDVIFGGGTKLTVL GQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASS YLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

Amino Acid Sequence Defining the PGT128 Derived Heavy Chain of the128/MV1 Antibody—PGT128-HC-Knob (SEQ ID NO:8):

QPQLQESGPTLVEASETLSLTCAVSGDSTAACNSF WGWVRQPPGKGLEWVGSLSHCASYWNRGWTYHNPSLKSRLTLALDTPKNLVFLKLNSVTAADTATYYCAR FGGEVLRYTDWPKPAWVDLWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVS WNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYN STYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWC LVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYT QKSLSLSPGK

10E8/MV1 Antibody:

Amino Acid Sequence Defining the MV1 Derived Light Chain of the 10E8/MV1Antibody—MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the 10E8/MV1Antibody—MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the 10E8 Derived Light Chain of the10E8/MV1 Antibody—10E8-LC (SEQ ID NO:9):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQ KKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKL TVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYA ASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8 Derived Heavy Chain of the10E8/MV1 Antibody—10E8-HC-Knob (SEQ ID NO:10):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMT WVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYY DFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPA PEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYR VVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

Δ10E8/MV1 Antibody

Amino Acid Sequence Defining the MV1 Derived Light Chain of theΔ10E8/MV1 Antibody MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of theΔ10E8/MV1 Antibody MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the Δ10E8 Derived Light Chain of theΔ10E8/MV1 Antibody Δ10E8-LC (SEQ ID NO:21):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGASFNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEG STVEKTVAPTECS

Amino Acid Sequence Defining the Δ10E8 Derived Heavy Chain of theΔ10E8/MV1 Antibody 10E8-HC-Knob (SEQ ID NO:22):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMTWVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYYDFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK

151/MV1 Antibody

Amino Acid Sequence Defining the MV1 Derived Light Chain of the 151/MV1Antibody—MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the 151/MV1Antibody MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHKANTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNAKLPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

Amino Acid Sequence Defining the PGT151 Derived Light Chain of the151/MV1 Antibody PGT151-LC (SEQ ID NO:23):

DIVMTQTPLSLSVTPGQPASISCKSSESLRQSNGKTSLYWYRQKPGQSPQLLVFEVSNRFSGVSDRFVGSGSGTDFTLRISRVEAEDVGFYYCMQSKDFPLTFGGGTKVDLKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACE VTHQGLSSPVTKSFNRGEC

Amino Acid Sequence Defining the PGT151 Derived Heavy Chain of the151/MV1 Antibody PGT151-HC-Knob (SEQ ID NO:24):

RVQLVESGGGVVQPGKSVRLSCVVSDFPFSKYPMYWVRQAPGKGLEWVAAISGDAWHVVYSNSVQGRFLVSRDNVKNTLYLEMNSLKIEDTAVYRCARMFQESGPPRLDRWSGRNYYYYSGMDVWGQGTIVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEAL HSHYTQKSLSLSPGK

145/P140 Antibody:

Amino Acid Sequence Defining the Pro 140 Derived Light Chain of the145/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the Pro 140 Derived Heavy Chain of the145/P140 Antibody—PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

Amino Acid Sequence Defining the PGT145 Derived Light Chain of the145/P140 Antibody—PGT145-LC (SEQ ID NO:3):

EVVITQSPLFLPVTPGEAASLSCKCSHSLQHSTGANYLATNYLQRPGQTPRLLIHLATHRASGVPDRFSGSGSGTDFTLKISRVESDDVGTYYCMQGLHSPTNTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQTNKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

Amino Acid Sequence Defining the PGT145 Derived Heavy Chain of the145/P140 Antibody—PGT145-HC-Knob (SEQ ID NO:4):

QVQLVQSGAEVKKPGSSVKVSCKASGNSFSNHDVHWVRQAPGQGLEWMGWMSHEGDKTGALQKFQGRVTITRDSGASTVYMELRGLTADDTAIYYCLTGSKHRLRDYFLYNEYGPNYEEWGDYLATLDVWGHGTAVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSV LHEALHSHYTQKSLSLSPGK

117/P140 Antibody:

Amino Acid Sequence Defining the Pro 140 Derived Light Chain of the117/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the Pro 140 Derived Heavy Chain of the117/P140 Antibody—PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

Amino Acid Sequence Defining the 3BNC117 Derived Light Chain of the117/P140 Antibody—3BNC117-LC (SEQ ID NO:5):

DIQMTQSPSSLSASVGDTVTITCQANGYLNWYQQRRGKAPKLLIYDGSKLERGVPSRFSGRRWGQEYNLTINNLQPEDIATYFCQVYEFVVFGQGTKVQVDIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSP VTKSFNRGEC

Amino Acid Sequence Defining the 3BNC117 Derived Heavy Chain of the117/P140 Antibody—3BNC117-HC-Knob (SEQ ID NO:6):

QVQLLQSGAAVTKPGASVEVSCEASGYNIRDYFIHWWRQAPGQGLQWVGWINPKTGQPNNPRQFQGRVSLTRHASWDFDTFSFYMDLKALRSDDTAVYFCARQRSDYWDFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLS PGK

128/P140 Antibody:

Amino Acid Sequence Defining the Pro 140 Derived Light Chain of the128/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the Pro 140 Derived Heavy Chain of the128/P140 Antibody—PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the PGT128 Derived Light Chain of the128/P140 Antibody—PGT128-LC (SEQ ID NO:7):

QSALTQPPSASGSPGQSITISCTGTSNNFVSWYQQHAGKAPKLVIYDVNKRPSGVPDRFSGSKSGNTASLTVSGLQTDDEAVYYCGSLVGNWDVIFGGGTKLTVLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKT VAPTECS

Amino Acid Sequence Defining the PGT128 Derived Heavy Chain of the128/P140 Antibody—PGT128-HC-Knob (SEQ ID NO:8):

QPQLQESGPTLVEASETLSLTCAVSGDSTAACNSFWGWVRQPPGKGLEWVGSLSHCASYWNRGWTYHNPSLKSRLTLALDTPKNLVFLKLNSVTAADTATYYCARFGGEVLRYTDWPKPAWVDLWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

10E8/P140 Antibody:

Amino Acid Sequence Defining the Pro 140 Derived Light Chain of the10E8/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the Pro 140 Derived Heavy Chain of the10E8/P140 Antibody—PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8 Derived Light Chain of the10E8/P140 Antibody—10E8-LC (SEQ ID NO:9):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTV EKTVAPTECS

Amino Acid Sequence Defining the 10E8 Derived Heavy Chain of the10E8/P140 Antibody—10E8-HC-Knob (SEQ ID NO:10):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMTWVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYYDFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

Δ10E8/P140 Antibody

Amino Acid Sequence Defining the PRO140 Derived Light Chain of theΔ10E8/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the PRO140 Derived Heavy Chain of theΔ10E8/P140 Antibody—PRO140-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the Δ10E8 Derived Light Chain of theΔ10E8/P140 Antibody—Δ10E8-LC (SEQ ID NO:21):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVILFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVE KTVAPTECS

Amino Acid Sequence Defining the Δ10E8 Derived Heavy Chain of theΔ10E8/P140 Antibody—10E8-HC-Knob (SEQ ID NO:22):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMTWVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYYDFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

151/P140 Antibody

Amino Acid Sequence Defining the PRO140 Derived Light Chain of the151/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the PRO140 Derived Heavy Chain of the151/P140 Antibody—PRO140-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the PGT151 Derived Light Chain of the151/P140 Antibody—PGT151-LC (SEQ ID NO:23):

DIVMTQTPLSLSVTPGQPASISCKSSESLRQSNGKTSLYWYRQKPGQSPQLLVFEVSNRFSGVSDRFVGSGSGTDFTLRISRVEAEDVGFYYCMQSKDFPLTFGGGTKVDLKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

Amino Acid Sequence Defining the PGT151 Derived Heavy Chain of the151/P140 Antibody—PGT151-HC-Knob (SEQ ID NO:24):

RVQLVESGGGVVQPGKSVRLSCVVSDFPFSKYPMYWVRQAPGKGLEWVAAISGDAWHVVYSNSVQGRFLVSRDNVKNTLYLEMNSLKIEDTAVYRCARMFQESGPPRLDRWSGRNYYYYSGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

10E8/Alpha-Her2 Antibody:

Amino Acid Sequence Defining the Alpha-Her2 Derived Light Chain of the10E8/Alpha-Her2 antibody—antiHer2-VLCH1 (SEQ ID NO:13):

DIVMTQSHKFMSTSVGDRVSITCKASQDVNTAVAWYQQKPGHSPKLLIYSASFRYTGVPDRFTGNRSGTDFTFTISSVQAEDLAVYYCQQHYTTPPTFGGGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSN TKVDKKVEPKSC 

Amino Acid Sequence Defining the Alpha-Her2 Derived Heavy Chain of the10E8/Alpha-Her2 Antibody—antiHer2-HC-Hole-Cross (SEQ ID NO:14):

QVQLQQSGPELVKPGASLKLSCTASGFNIKDTYIHWVKQRPEQGLEWIGRIYPTNGYTRYDPKFQDKATITADTSSNTAYLQVSRLTSEDTAVYYCSRWGGDGFYAMDYWGQGASVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFMNYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRTNQQGNVFSCSVLHEALHSHYTQKSLS LSPGK 

Amino Acid Sequence Defining the 10E8 Derived Light Chain of the10E8/Alpha-Her2 Antibody—10E8-LC (SEQ ID NO:9):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHE GSTVEKTVAPTECS 

Amino Acid Sequence Defining the 10E8 Derived Heavy Chain of the10E8/Alpha-Her2 Antibody—10E8-HC-Knob (SEQ ID NO:10):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMTWVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYYDFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK 

4E10/P140 Antibody:

Amino Acid Sequence Defining the Pro 140 Derived Light Chain of the4E10/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC 

Amino Acid Sequence Defining the Pro 140 Derived Heavy Chain of the4E10/P140 Antibody—PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK 

Amino Acid Sequence Defining the 4E10 Derived Light Chain of the4E10/P140 Antibody—4E10-LC (SEQ ID NO:17):

EIVLIQSPGTQSLSPGERATLSCRASQSVGNNKLAWYQQRPGQAPRLLIYGASSRPSGVADRFSGSGSGTDFTLTISRLEPEDFAVYYCQQYGQSLSTFGQGTKVEVKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSILTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC 

Amino Acid Sequence Defining the 4E10 Derived Heavy Chain of the4E10/P140 Antibody—PGT145-HC-Knob (SEQ ID NO:18):

QVQLVQSGAEVKRPGSSVTVSCKASGGSFSTYALSWVRQAPGRGLEWMGGVIPLLTITNYAPRFQGRITITADRSTSTAYLELNSLRPEDTAVYYCAREGTTGAGWLGKPIGAFAHWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTITPVLDSDGSFFLYSKLTVDKSRTA7QQGNVFSCSVLHEALHSHYTQ KSLSLSPGK 

10E8/X19 Antibody:

Amino Acid Sequence Defining the X19 Derived Light Chain of the 10E8/X19Antibody—X19-VLCH1 (SEQ ID NO:19):

EIVLIQSPATLSVSPGRRATLSCRASQSVNTNLAWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQHYGSSPLIFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSN TKVDKKVEPKSC 

Amino Acid Sequence Defining the X19 Derived Heavy Chain of the 10E8/X19Antibody—X19-HC-Hole-Cross (SEQ ID NO:20):

QVQLVQSGGGVVQPGRSLRLSCAASGFTFSSYPMHWVRQAPGKGLEWMTVISSDGRNKYYPDSVKGRFTISRDNSKNTLYLQMNSLRPEDTAVYYCARGGYHDFWSGPDYWGQGTLVIVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQNKVDNALQSGNSQESVTEQDSKDSTYSLSSILTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTITPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLS LSPGK 

Amino Acid Sequence Defining the 10E8 Derived Light Chain of the10E8/X19 Antibody—10E8-LC (SEQ ID NO:9):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHE GSTVEKTVAPTECS 

Amino Acid Sequence Defining the 10E8 Derived Heavy Chain of the10E8/X19 Antibody—PGT145-HC-Knob (SEQ ID NO:10):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMTWVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYYDFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK 

10E8/515H7 Antibody

Amino Acid Sequence Defining the 515H7 Derived Light Chain of the10E8/515H7 Antibody—515H7-VLCH1 (SEQ ID NO:25):

DIVMSQSPSSLAVSAGEKVTMSCKSSQSLFNSRTRKNYLAWYQQKPGQSPKLLIYWASARDSGVPARFTGSGSETYFTLTISRVQAEDLAVYYCMQSFNLRTFGGGTKLEIKASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK PSNTKVDKKVEPKSC 

Amino Acid Sequence Defining the 515H7 Derived Heavy Chain of the10E8/515H7 Antibody—515H7-Hole-Cross (SEQ ID NO:26):

EVNLVESGGGLVQPGGSLRLSCATSGFTFTDNYMSWVRQPPGKALEWLGFIRNKANGYTTDYSASVRGRFTISRDNSQSILYLQMNALRAEDSATYYCARDVGSNYFDYWGQGTTLIVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRTNQQGNVFSCSVLHEALHSHYTQKSLS LSPGK 

Amino Acid Sequence Defining the 10E8 Derived Light Chain of the10E8/515H7 Antibody—10E8-LC (SEQ ID NO:9):

YELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHE GSTVEKTVAPTECS 

Amino Acid Sequence Defining the 10E8 Derived Heavy Chain of the10E8/515H7 Antibody—10E8-HC-Knob (SEQ ID NO:10):

EVQLVESGGGLVKPGGSLRLSCSASGFDFDNAWMTWVRQPPGKGLEWVGRITGPGEGWSVDYAAPVEGRFTISRLNSINFLYLEMNNLRMEDSGLYFCARTGKYYDFWSGYPPGEEYFQDWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

Chimeric CDR123 Antibody (SEQ ID NO:27):

SELTQDPAVSVALGQTVRITCRGDSLRSHYASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTV EKTVAPTECS

Chimeric FW123 (SEQ ID NO:28):

YELTQETGVSVALGRTVTITCQGDSLRSYYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCNSRDSSGNHLVVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTV EKTVAPTECS

10E8V1.0/iMab Antibody

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8v1.0/MV1 Antibody MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS KNTKVDKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8v1.0/MV1 Antibody MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8v1.0 Derived Light Chain of the10E8v1.0/iMab Antibody—10E8v1.0-LC (SEQ ID NO:29):

ASELTQDPAVSVALKQTVTITCRGDSLRSHYVSWYQKKPGQAPVLVFYGKNNRPSGIPDRFSGSSSGNTASLTIAGAQAEDDADYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGST VEKTVAPTECS

Amino Acid Sequence Defining the 10E8v1.0 Derived Heavy Chain of the10E8v1.0/iMab Antibody—10E8v1.0-HC-Knob (SEQ ID NO:30):

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

10E8V1.1/iMab Antibody

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8v1.1/iMab Antibody MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8v1.1/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8v1.1 Derived Light Chain of the10E8v1.1/iMab Antibody—10E8v1.1-LC (SEQ ID NO:31):

ASELTQDPAVSVALKQTVTITCRGDSLRSHYVSWYQKKPGQAPVLVFYGKNNRPSGIPDRFSGSSSGNTASLTIAGAQAEDDADYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQNKSHRSYSCQVTHEGST VEKTVAPTECS

Amino Acid Sequence Defining the 10E8v1.1 Derived Heavy Chain of the10E8v1.1/iMab Antibody—10E8v1.1 HC-Knob (SEQ ID NO:32):

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKYYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

10E8V2.0/iMab Antibody (Also Referred to as 10E8.2/iMab Antibody)

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8v2.0/iMab Antibody MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8v2.0/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFMNYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8v2.0 Derived Light Chain of the10E8v2.0/iMab Antibody—10E8v2.0-LC (SEQ ID NO:33):

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLEPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGST VEKTVAPTECS

Amino Acid Sequence Defining the 10E8v2.0 Derived Heavy Chain of the10E8v2.0/iMab Antibody—10E8v2.0-HC-Knob (SEQ ID NO:34):

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

10E8V3.0/iMab Antibody

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8v3.0/iMab Antibody MV1-VLCH1 (SEQ ID NO:1):

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8v3.0/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO:2):

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8v3.0 Derived Light Chain of the10E8v3.0/iMab Antibody—10E8v3.0-LC (SEQ ID NO:15):

SELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGIHDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTV EKTVAPTECS

Amino Acid Sequence Defining the 10E8v3.0 Derived Heavy Chain of the10E8v3.0/iMab Antibody—10E8v3.0-HC-Knob (SEQ ID NO:16):

EVQLVESGGDLVKPGGSLRLSCSASGFSFKNTWMTWVRQAPGKGLEWVGRITGPGEGWTSDYAATVQGRFTISRNNMIDMLYLEMNRLRTDDTGLYYCVHTEKYYNFWGGYPPGEEYFQHWGRGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

10E8V1.0/P140 (H6L10/PRO140) Antibody

Amino Acid Sequence Defining the PRO140 Derived Light Chain of the10E8V1.0/P140 Antibody—PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the PRO140 Derived Heavy Chain of the10E8V1.0/P140 Antibody—PRO140-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the L10 Derived Light Chain of the10E8V1.0/P140 Antibody—L10-LC (SEQ ID NO:29):

ASELTQDPAVSVALKQTVTITCRGDSLRSHYVSWYQKKPGQAPVLVFYGKNNRPSGIPDRFSGSSSGNTASLTIAGAQAEDDADYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGST VEKTVAPTECS

Amino Acid Sequence Defining the H6 Derived Heavy Chain of the10E8V1.0/P140 Antibody—H6-HC-Knob (SEQ ID NO:30):

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

10E8V1.1/P140 Antibody

Amino Acid Sequence Defining the PRO140 Derived Light Chain of the10E8v1.1/P140 Antibody PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the P140 Derived Heavy Chain of the10E8v1.1/P140 Antibody PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8v1.1 Derived Light Chain of the10E8v1.1/P140 Antibody—10E8v1.1-LC (SEQ ID NO:31):

ASELTQDPAVSVALKQTVTITCRGDSLRSHYVSWYQKKPGQAPVLVFYGKNNRPSGIPDRFSGSSSGNTASLTIAGAQAEDDADYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLEPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGST VEKTVAPTECS

Amino Acid Sequence Defining the 10E8v1.1 Derived Heavy Chain of the10E8v1.1/P140 Antibody—10E8v1.1 HC-Knob (SEQ ID NO:32):

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKYYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSP GK

10E8V2.0/P140 Antibody

Amino Acid Sequence Defining the PRO140 Derived Light Chain of the10E8v2.0/P140 Antibody PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS NTKVDKKVEPKSC

Amino Acid Sequence Defining the P140 Derived Heavy Chain of the10E8v2.0/P140 Antibody PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK

Amino Acid Sequence Defining the 10E8v2.0 Derived Light Chain of the10E8v2.0/P140 Antibody—10E8v2.0-LC (SEQ ID NO:33):

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGST VEKTVAPTECS

Amino Acid Sequence Defining the 10E8v2.0 Derived Heavy Chain of the10E8v2.0/P140 Antibody—10E8v2.0 HC-Knob (SEQ ID NO:34):

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK 

10E8V3.0/P140 Antibody

Amino Acid Sequence Defining the PRO140 Derived Light Chain of the10E8v3.0/P140 Antibody PRO140-VLCH1 (SEQ ID NO:11):

DIVMTQSPLSLPVTPGEPASISCRSSQRLLSSYGHTYLHWYLQKPGQSPQLLIYEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPLTFGQGTKVEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC 

Amino Acid Sequence Defining the P140 Derived Heavy Chain of the10E8v3.0/P140 Antibody PRO140-HC-Hole-Cross (SEQ ID NO:12):

EVQLVESGGGLVKPGGSLRLSCAASGYTFSNYWIGWVRQAPGKGLEWIGDIYPGGNYIRNNEKFKDKTTLSADTSKNTAYLQMNSLKTEDTAVYYCGSSFGSNYVFAWFTYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK 

Amino Acid Sequence Defining the 10E8v3.0 Derived Light Chain of the10E8v3.0/P140 Antibody—10E8v3.0-LC (SEQ ID NO:15):

SELTQETGVSVALGRTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGIHDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHE GSTVEKTVAPTECS 

Amino Acid Sequence Defining the 10E8v3.0 Derived Heavy Chain of the10E8v3.0/P140 Antibody—10E8v3.0 HC-Knob (SEQ ID NO:16):

EVQLVESGGDLVKPGGSLRLSCSASGFSFKNTWMTWVRQAPGKGLEWVGRITGPGEGWTSDYAATVQGRFTISRNNMIDMLYLEMNRLRTDDTGLYYCVHTEKYYNFWGGYPPGEEYFQHWGRGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK 

10E8.2.1/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC 

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFMNYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK 

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLEPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEONKSHRSYSCQVTH EGSTVEKTVAPTECS 

Amino Acid Sequence Defining the 10E8.2.1 Derived Heavy Chain of the10E8.2.1/iMab Antibody—10E8.2.1-HC-Knob (SEQ ID NO:35)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRDNTKNTLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTLVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK 

10E8.2.2/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC 

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRTNQQGNVFSCSVLHEALHSHYTQKS LSLSPGK 

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVEGGGTKLTVLSQPKAAPSVTLEPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEONKSHRSYSCQVTH EGSTVEKTVAPTECS 

Amino Acid Sequence Defining the 10E8.2.2 Derived Heavy Chain of the10E8.2.2/iMab Antibody—10E8.2.2-HC-Knob (SEQ ID NO:36)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRLNSINFLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK 

10E8.2.3/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC 

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVICVVVDVSHEDPEVKFMNYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRTNQQGNVFSCSVLHEALHSHYTQKS LSLSPGK 

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTH EGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.2.3 Derived Heavy Chain of the10E8.2.3/iMab Antibody—10E8.2.3-HC-Knob (SEQ ID NO:37)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRDNTKNTLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK

10E8.2.4/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2.4/iMab Antibody MV1-VLCH1-LM52 (SEQ ID NO:38)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWANSTESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQEPNNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGVNFSCSVLHEALHSHYTQKSL SLSPGK

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTH EGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.2.3 Derived Heavy Chain of the10E8.2.3/iMab Antibody—10E8.2.3-HC-Knob (SEQ ID NO:39)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRDNTKNTLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPECTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK

10E8.2.5/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIHWVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTH EGSTVEVTVAPTECS

Amino Acid Sequence Defining the 10E8.2.5 Derived Heavy Chain of the10E8.2.5/iMab Antibody—10E8.2.5-HC-Knob (SEQ ID NO:40)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRKNSINTLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK

10E8.2.6/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIWHVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYATGAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNAKLPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWYQKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTKLTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTH EGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.2.6 Derived Heavy Chain of the10E8.2.6/iMab Antibody—10E8.2.6-HC-Knob (SEQ ID NO:41)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMTWVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRDNSINTLYLEMNNVRTEDTGYYFCARTGKHYDFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPECTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHY TQKSLSLSPGK

10E8.2.7/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQKNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGGGTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN HKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWOOGNVFSCSVLHEALHSHYTOKSLSLSPG K 

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWY QKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTK LTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKY AASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.4 Derived Heavy Chain of the10E8.4/iMab Antibody—10E8.4-HC-Knob (SEQ ID NO:42)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMT WVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRKNSKNTLYLEMNNVRTEDTGYYFCARTGKHY DFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPA PEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYR VVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

10E8.2.8/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the 10E8.2 Derived Light Chain of the10E8.2/iMab Antibody—10E8.2-LC (SEQ ID NO: 33)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWY QKKPGQAPILLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTK LTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKY AASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.2.8 Derived Heavy Chain of the10E8.2.8/iMab Antibody—10E8.2.8-HC-Knob (SEQ ID NO:43)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMT WVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRDNSKNTLYLEMNNVRTEDTGYYFCARTGKHY DFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPA PEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYR VVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

10E8.2.10/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the 10E8.4 Derived Light Chain of the10E8.4/iMab Antibody—10E8.4-LC (SEQ ID NO:44)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWY QKKTGQAPKLLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTK LTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKY AASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.2.8 Derived Heavy Chain of the10E8.2.8/iMab Antibody—10E8.2.8-HC-Knob (SEQ ID NO:45)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMT WVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRDNSKNTLYLEMNNVRTEDTGYYFCARTGKHY DFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPA PEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYR VVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

10E8.4/iMab

Amino Acid Sequence Defining the MV1 Derived Light Chain of the10E8.2/iMab Antibody MV1-VLCH1 (SEQ ID NO: 1)

DIVMTQSPDSLAVSLGERVTMNCKSSQSLLYSTNQ KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSVQAEDVAVYYCQQYYSYRTFGG GTKLEIKSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC

Amino Acid Sequence Defining the MV1 Derived Heavy Chain of the10E8.2/iMab Antibody MV1-HC-Hole-Cross (SEQ ID NO: 2)

QVQLQQSGPEVVKPGASVKMSCKASGYTFTSYVIH WVRQKPGQGLDWIGYINPYNDGTDYDEKFKGKATLTSDTSTSTAYMELSSLRSEDTAVYYCAREKDNYAT GAWFAYWGQGTLVTVSSASTAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQ ESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECDKTHTCPPCPAPEFEG GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVL TVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPG K

Amino Acid Sequence Defining the 10E8.4 Derived Light Chain of the10E8.4/iMab Antibody—10E8.4-LC (SEQ ID NO:46)

ASELTQDPAVSVALKQTVTITCRGDSLRSHYASWY QKKTGQAPKLLFYGKNNRPSGVPDRFSGSASGNRASLTISGAQAEDDAEYYCSSRDKSGSRLSVFGGGTK LTVLSQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKY AASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

Amino Acid Sequence Defining the 10E8.4 Derived Heavy Chain of the10E8.4/iMab Antibody—10E8.4-HC-Knob (SEQ ID NO:47)

EVRLVESGGGLVKPGGSLRLSCSASGFNFDDAWMT WVRQPPGKGLEWVGRISGPGEGWSVDYAESVKGRFTISRKNSKNTLYLEMNNVRTEDTGYYFCARTGKHY DFWSGYPPGEEYFQDWGQGTKVIVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPA PEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYR VVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVLHEALHSHYTQKSL SLSPGK

In various embodiments, at least one of the heavy chain and/or lightchain sequences derived from the PGT145, PG9, PGT128, PGT121, 10-1074,3BNC117, VRC01, PGT151, 4E10, 10E8, P140, iMab (or the MV1 variant),515H7 antibodies and variants thereof are paired together to form abispecific antibody (e.g., a HIV CrossMab antibody). In an exemplaryembodiment, at least one of the disclosed heavy and light chainsselected from SEQ ID NOs: 1-36 are paired together to form a bispecificantibody (e.g., a HIV CrossMab antibody).

In various embodiments, the amino acid sequence of the bispecificantibody (e.g., HIV CrossMab antibody) further includes an amino acidanalog, an amino acid derivative, or other non-classical amino acids.

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that is at least 60% identical to awild-type heavy or light chain sequence of the PGT145, PG9, PGT128,PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10, or 10E8 antibody. Invarious embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that is at least 60% identical to awild-type heavy chain or light chain sequence of the P140, iMab (or theMV1 variant), or 515H7 antibody. In exemplary embodiments, thebispecific antibody (e.g., HIV CrossMab antibody) comprises a sequencethat is at least 60% identical to any of the sequences disclosed herein.

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) may comprise a sequence that is at least about 60%, at leastabout 61%, at least about 62%, at least about 63%, at least about 64%,at least about 65%, at least about 66%, at least about 67%, at leastabout 68%, at least about 69%, at least about 70%, at least about 71%,at least about 72%, at least about 73%, at least about 74%, at leastabout 75%, at least about 76%, at least about 77%, at least about 78%,at least about 79%, at least about 80%, at least about 81%, at leastabout 82%, at least about 83%, at least about 84%, at least about 85%,at least about 86%, at least about 87%, at least about 88%, at leastabout 89%, at least about 90%, at least about 91%, at least about 92%,at least about 93%, at least about 94%, at least about 95%, at leastabout 96%, at least about 97%, at least about 98%, at least about 99%,or 100% identical to a wild-type heavy chain or light chain sequence ofthe PGT145, PG9, PGT128, PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10,or 10E8 antibody.

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) may comprise a sequence that is at least about 60%, at leastabout 61%, at least about 62%, at least about 63%, at least about 64%,at least about 65%, at least about 66%, at least about 67%, at leastabout 68%, at least about 69%, at least about 70%, at least about 71%,at least about 72%, at least about 73%, at least about 74%, at leastabout 75%, at least about 76%, at least about 77%, at least about 78%,at least about 79%, at least about 80%, at least about 81%, at leastabout 82%, at least about 83%, at least about 84%, at least about 85%,at least about 86%, at least about 87%, at least about 88%, at leastabout 89%, at least about 90%, at least about 91%, at least about 92%,at least about 93%, at least about 94%, at least about 95%, at leastabout 96%, at least about 97%, at least about 98%, at least about 99%,or 100% identical to a wild-type heavy chain or light chain sequence ofthe P140, iMab (or the MV1 variant), or 515H7 antibody.

In exemplary embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) may comprise a sequence that is at least about 60%, at leastabout 61%, at least about 62%, at least about 63%, at least about 64%,at least about 65%, at least about 66%, at least about 67%, at leastabout 68%, at least about 69%, at least about 70%, at least about 71%,at least about 72%, at least about 73%, at least about 74%, at leastabout 75%, at least about 76%, at least about 77%, at least about 78%,at least about 79%, at least about 80%, at least about 81%, at leastabout 82%, at least about 83%, at least about 84%, at least about 85%,at least about 86%, at least about 87%, at least about 88%, at leastabout 89%, at least about 90%, at least about 91%, at least about 92%,at least about 93%, at least about 94%, at least about 95%, at leastabout 96%, at least about 97%, at least about 98%, at least about 99%,or 100% identical to any of the sequences disclosed herein.

Homology or identity may be determined in various ways that are withinthe skill in the art, for instance, using publicly available computersoftware such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software.BLAST (Basic Local Alignment Search Tool) analysis using the algorithmemployed by the programs blastp, blastn, blastx, tblastn and tblastx(Karlin et al., (1990) PROC. NATL. ACAD. SCI. USA 87, 2264-2268;Altschul, (1993) J. MOL. EVOL. 36, 290-300; Altschul et al., (1997)NUCLEIC ACIDS RES. 25, 3389-3402, incorporated by reference) aretailored for sequence similarity searching. The approach used by theBLAST program is to first consider similar segments between a querysequence and a database sequence, then to evaluate the statisticalsignificance of all matches that are identified and finally to summarizeonly those matches which satisfy a preselected threshold ofsignificance. For a discussion of basic issues in similarity searchingof sequence databases see Altschul et al., (1994) NATURE GENETICS 6,119-129 which is fully incorporated by reference. Those skilled in theart can determine appropriate parameters for measuring alignment,including any algorithms needed to achieve maximal alignment over thefull length of the sequences being compared. The search parameters forhistogram, descriptions, alignments, expect (i.e., the statisticalsignificance threshold for reporting matches against databasesequences), cutoff, matrix and filter are at the default settings. Thedefault scoring matrix used by blastp, blastx, tblastn, and tblastx isthe BLOSUM62 matrix (Henikoff et al., (1992) PROC. NATL. ACAD. SCI. USA89, 10915-10919, fully incorporated by reference). Four blastnparameters may be adjusted as follows: Q=10 (gap creation penalty); R=10(gap extension penalty); wink=1 (generates word hits at everywink.sup.th position along the query); and gapw=16 (sets the windowwidth within which gapped alignments are generated). The equivalentBlastp parameter settings may be Q=9; R=2; wink=1; and gapw=32. Searchesmay also be conducted using the NCBI (National Center for BiotechnologyInformation) BLAST Advanced Option parameter (e.g.: −G, Cost to open gap[Integer]: default=5 for nucleotides/11 for proteins; −E, Cost to extendgap [Integer]: default=2 for nucleotides/1 for proteins; −q, Penalty fornucleotide mismatch [Integer]: default=−3; −r, reward for nucleotidematch [Integer]: default=1; −e, expect value [Real]: default=10; −W,wordsize [Integer]: default=11 for nucleotides/28 for megablast/3 forproteins; −y, Dropoff (X) for blast extensions in bits: default=20 forblastn/7 for others; −X, X dropoff value for gapped alignment (in bits):default=15 for all programs, not applicable to blastn; and —Z, final Xdropoff value for gapped alignment (in bits): 50 for blastn, 25 forothers). ClustalW for pairwise protein alignments may also be used(default parameters may include, e.g., Blosum62 matrix and Gap OpeningPenalty=10 and Gap Extension Penalty=0.1). A Bestfit comparison betweensequences, available in the GCG package version 10.0, uses DNAparameters GAP=50 (gap creation penalty) and LEN=3 (gap extensionpenalty) and the equivalent settings in protein comparisons are GAP=8and LEN=2.

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that includes at least one amino acidalteration with respect to a wild-type heavy or light chain sequence ofthe PGT145, PG9, PGT128, PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10,or 10E8 antibody. In various embodiments, the bispecific antibody (e.g.,HIV CrossMab antibody) comprises a sequence that includes at least oneamino acid alteration with respect to a wild-type heavy or light chainsequence of the P140, iMab (or the MV1 variant), 515H7 antibody. Inexemplary embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that includes at least one amino acidalteration with respect to any of the sequences disclosed herein.

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that includes at least about 1, 2, 3, 4,5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23,24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41,42, 43, 44, 45, 46, 47, 40, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77,78, 79, or 80 amino acid alterations with respect to a wild-type heavyor light chain sequence of the PGT145, PG9, PGT128, PGT121, 10-1074,3BNC117, VRC01, PGT151, 4E10, or 10E8 antibody.

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that includes at least about 1, 2, 3, 4,5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23,24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41,42, 43, 44, 45, 46, 47, 40, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77,78, 79, or 80 amino acid alterations with respect to a wild-type heavyor light chain sequence of the P140, iMab (or the MV1 variant), or 515H7antibody.

In exemplary embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) comprises a sequence that includes at least about 1, 2, 3, 4,5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23,24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41,42, 43, 44, 45, 46, 47, 40, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77,78, 79, or 80 amino acid alterations with respect to any of thesequences disclosed herein.

The amino acid alteration can be an amino acid deletion, insertion,substitution, or modification. In one embodiment, the amino acidalteration is an amino acid deletion. In another embodiment, the aminoacid alteration is an amino acid substitution.

In various embodiments, the amino acid alteration may be in theComplementarity Determining Regions (CDRs) of the bispecific antibody(e.g., the CDR1, CDR2 or CDR3 regions). In another embodiment, the aminoacid alteration may be in the framework regions (FWs) of the bispecificantibody (e.g., the FW1, FW2, FW3, or FW4 regions). In a furtherembodiment, the amino acid alteration may be in the joining regions (Jregions) of the bispecific antibody (e.g., the J1, J2, J3, J4, J5, J6,or J7 regions).

Also provided herein are chimeric antibody derivatives of the bispecificantibodies, i.e., antibody molecules in which a portion of the heavyand/or light chain is identical with or homologous to correspondingsequences in antibodies derived from a particular species or belongingto a particular antibody class or subclass, while the remainder of thechain(s) is identical with or homologous to corresponding sequences inantibodies derived from another species or belonging to another antibodyclass or subclass, as well as fragments of such antibodies, so long asthey exhibit the desired biological activity. For example, thebispecific antibody may include a heavy and/or light chain in which oneor more CDRs or FWs derived from an antibody selected from a PGT145,PG9, PGT128, PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10, 10E8, P140,iMab (or the MV1 variant), or 515H7 antibody are replaced with one ormore CDRs or FWs derived from a different antibody selected from aPGT145, PG9, PGT128, PGT121, 10-1074, 3BNC117, VRC01, PGT151, 4E10,10E8, P140, iMab (or the MV1 variant), or 515H7 antibody.

In various embodiments, the present invention provides improvedbispecific antibodies that demonstrate advantageous properties relatedto solubility, stability, and therapeutic activity. It is contemplatedthat such antibodies may be particularly suited for large-scalecommercial production. For example, such antibodies may exhibitincreased solubility, reduced aggregation, reduced precipitation, and/orenhanced stability or resistance to degradation during manufacturing.

In an exemplary embodiment, the improved bispecific antibody is avariant of the 10E8V2.0/iMab antibody (also referred to as 10E8.2/iMabantibody). In such embodiments, the variant may exhibit enhancedsolubility, stability, and/or therapeutic activity (e.g., antiviralactivity) compared to the parent 10E8V2.0/iMab antibody.

In some embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) may comprise a sequence that is at least about 60%, at leastabout 61%, at least about 62%, at least about 63%, at least about 64%,at least about 65%, at least about 66%, at least about 67%, at leastabout 68%, at least about 69%, at least about 70%, at least about 71%,at least about 72%, at least about 73%, at least about 74%, at leastabout 75%, at least about 76%, at least about 77%, at least about 78%,at least about 79%, at least about 80%, at least about 81%, at leastabout 82%, at least about 83%, at least about 84%, at least about 85%,at least about 86%, at least about 87%, at least about 88%, at leastabout 89%, at least about 90%, at least about 91%, at least about 92%,at least about 93%, at least about 94%, at least about 95%, at leastabout 96%, at least about 97%, at least about 98%, at least about 99%,or 100% identical to the heavy chain or light chain sequences of the10E8V2.0/iMab antibody (i.e., SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO: 33,or SEQ ID NO:34).

In various embodiments, the bispecific antibody (e.g., HIV CrossMabantibody) may comprise a sequence that includes at least about 1, 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22,23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40,41, 42, 43, 44, 45, 46, 47, 40, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58,59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76,77, 78, 79, or 80 amino acid alterations with respect to the heavy chainor light chain sequences of the 10E8V2.0/iMab antibody (i.e., SEQ IDNO:1, SEQ ID NO:2, SEQ ID NO: 33, or SEQ ID NO:34).

In various embodiments, the bispecific antibody may comprise one or moreamino acid alterations in the Complementarity Determining Regions (CDRs)of the 10E8V2.0/iMab antibody (e.g., the CDR1, CDR2 or CDR3 regions). Inanother embodiment, the bispecific antibody may comprise one or moreamino acid alterations in the framework regions (FWs) of the bispecificantibody (e.g., the FW1, FW2, FW3, or FW4 regions). In a furtherembodiment, the amino acid alterations may be in the joining regions (Jregions) of the 10E8V2.0/iMab antibody (e.g., the J1, J2, J3, J4, J5,J6, or J7 regions).

In some embodiments, the bispecific antibody comprises a variant heavychain derived from 10E8V2.0 (i.e., SEQ ID NO:34). In such embodiments,the bispecific antibody may include one or more mutations at positionsselected from L72, I75, F77, L89, Y98, F100a, W100b, Y100e, P100f,P100g, L108, and/or L170 of the heavy chain (the mutation positions onSEQ ID NO: 34 are determined by the Kabat numbering system). In someembodiments, the bispecific antibody may include one or more mutationsat positions selected from L72, I75, F77, and/or L108. In someembodiment, the bispecific antibody may include the one or moremutations selected from L72K, I75K, F77T, and L108K. In an embodiment,the bispecific antibody comprises a heavy chain comprising the aminoacid sequence of SEQ ID NO: 47.

In some embodiments, the bispecific antibody comprises a variant lightchain derived from 10E8V2.0 (i.e., SEQ ID NO:33). In such embodiments,the bispecific antibody may include one or more mutations at positionsselected from L15, P40, I45, and P112 of the light chain (the mutationpositions on SEQ ID NO: 33 are determined by the Kabat numberingsystem). In some embodiments, the bispecific antibody may include one ormore mutations at positions selected from P40 and I45. In someembodiments, the bispecific antibody may include one or more mutationsselected from P4OT and I45K. In an embodiment, the bispecific antibodycomprises a light chain comprising the amino acid sequence of SEQ ID NO:46.

In some embodiments, the bispecific antibody comprises a variant lightchain derived from MV1 (i.e., SEQ ID NO: 1). In such embodiments, thebispecific antibody may include mutations at positions 52-54 (themutation position on SEQ ID NO: 1 is determined by the Kabat numberingsystem). In some embodiments, the bispecific antibody may include aminoacid mutations at positions 52-54. In some embodiments, the amino acidat position 52 is mutated to Asn (N), the amino acid at position 53 ismutated to Ser (S), and the amino acid at position 54 is mutated to Thr(T). In some embodiments, the Asn mutation at position 52 is N-linkedglycosylated. In an embodiment, the bispecific antibody comprises alight chain comprising the amino acid sequence of SEQ ID NO: 38.

In illustrative embodiment, the bispecific antibody comprises a heavyand light chain derived from 10E8 comprising the amino acid sequence ofSEQ ID NO: 47 and SEQ ID NO:46, respectively. The bispecific antibodyfurther comprises a heavy and light chain derived from MV1 comprisingthe amino acid sequence of SEQ ID NO: 1 and SEQ ID NO:2, respectively.

Modification of the amino acid sequence of recombinant binding proteinis achieved using any known technique in the art e.g., site-directedmutagenesis or PCR based mutagenesis. Such techniques are described, forexample, in Sambrook et al., Molecular Cloning: A Laboratory Manual,Cold Spring Harbor Press, Plainview, N.Y., 1989 and Ausubel et al.,Current Protocols in Molecular Biology, John Wiley & Sons, New York,N.Y., 1989.

Methods for producing antibodies, such as those disclosed herein, areknown in the art. For example, DNA molecules encoding light chainvariable regions and/or heavy chain variable regions can be chemicallysynthesized using the sequence information provided herein. SyntheticDNA molecules can be ligated to other appropriate nucleotide sequences,including, e.g., expression control sequences, to produce conventionalgene expression constructs encoding the desired antibodies. Productionof defined gene constructs is within routine skill in the art.Alternatively, the sequences provided herein can be cloned out ofhybridomas by conventional hybridization techniques or polymerase chainreaction (PCR) techniques, using synthetic nucleic acid probes whosesequences are based on sequence information provided herein, or priorart sequence information regarding genes encoding the heavy and lightchains.

Nucleic acids encoding desired antibodies can be incorporated (ligated)into expression vectors, which can be introduced into host cells throughconventional transfection or transformation techniques. Exemplary hostcells are E. coli cells, Chinese hamster ovary (CHO) cells, humanembryonic kidney 293 (HEK 293) cells, HeLa cells, baby hamster kidney(BHK) cells, monkey kidney cells (COS), human hepatocellular carcinomacells (e.g., Hep G2), and myeloma cells that do not otherwise produceIgG protein. Transformed host cells can be grown under conditions thatpermit the host cells to express the genes that encode theimmunoglobulin light and/or heavy chain variable regions. Specificexpression and purification conditions will vary depending upon theexpression system employed.

In various embodiments, the bispecific antibodies of the presentinvention (e.g., HIV CrossMab antibodies) are used in therapy. Forexample, the bispecific antibody (e.g., HIV CrossMab antibody) can beused to neutralize HIV in a mammal (e.g., a human patient). For example,antibodies of the invention can bind to HIV so as to partially orcompletely inhibit one or more biological activities of the virus. In anembodiment, the bispecific antibody (e.g., HIV CrossMab antibody)neutralizes a R5-tropic HIV. In another embodiment, the bispecificantibody (e.g., HIV CrossMab antibody) neutralizes a X4-tropic HIV. In afurther embodiment, the bispecific antibody (e.g., HIV CrossMabantibody) neutralizes a R5X4 dual-tropic HIV. In some embodiments, useof the antibody to neutralize HIV in a mammal comprises administering tothe mammal a therapeutically effective amount of the antibody.

Generally, a therapeutically effective amount of active component is inthe range of, for example, about 0.1 mg/kg to about 100 mg/kg, e.g.,about 1 mg/kg to about 100 mg/kg, e.g., about 1 mg/kg to about 10 mg/kgof the body weight of the patient. In various embodiments, atherapeutically effective amount of active component is in a range ofabout 0.01 mg/kg to about 30 mg/kg of the body weight of the patient,for example, about 0.01 mg/kg, about 0.02 mg/kg, about 0.03 mg/kg, about0.04 mg/kg, about 0.05 mg/kg, about 0.06 mg/kg, about 0.07 mg/kg, about0.08 mg/kg, about 0.09 mg/kg, about 0.1 mg/kg, about 0.2 mg/kg, about0.3 mg/kg, about 0.4 mg/kg, about 0.5 mg/kg, about 0.6 mg/kg, about 0.7mg/kg, about 0.8 mg/kg, about 0.9 mg/kg, about 1 mg/kg, about 1.1 mg/kg,about 1.2 mg/kg, about 1.3 mg/kg, about 1.4 mg/kg, about 1.5 mg/kg,about 1.6 mg/kg, about 1.7 mg/kg, about 1.8 mg/kg, 1.9 mg/kg, about 2mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about7 mg/kg, about 8 mg/kg, about 9 mg/kg, about 10 mg/kg, about 11 mg/kg,about 12 mg/kg, about 13 mg/kg, about 14 mg/kg, about 15 mg/kg, about 16mg/kg, about 17 mg/kg, about 18 mg/kg, about 19 mg/kg, about 20 mg/kg,about 21 mg/kg, about 22 mg/kg, about 23 mg/kg, about 24 mg/kg, about 25mg/kg, about 26 mg/kg, about 27 mg/kg, about 28 mg/kg, about 29 mg/kg,about 30 mg/kg body weight, inclusive of all values and ranges therebetween.

In some embodiments, the therapeutically effective amount of activecomponent is any value between about 1 to 10 mg/kg, between about 10 to20 mg/kg, between about 20 to 30 mg/kg, between about 30 to 40 mg/kg,between about 40 to 50 mg/kg, between about 50 to 60 mg/kg, betweenabout 60 to 70 mg/kg, between about 70 to 80 mg/kg, between about 80 to90 mg/kg, or between about 90 to 100 mg/kg.

In some embodiments, the therapeutically effective amount of activecomponent is about 1 mg/kg, about 2 mg/kg, about 3 mg/kg, about 6 mg/kg,about 10 mg/kg, about 20 mg/kg, about 30 mg/kg, or about 60 mg/kgdelivered intravenously (IV).

In some embodiments, the therapeutically effective amount of activecomponent is about 2.5 mg/kg, about 5 mg/kg, about 10 mg/kg, or about 20mg/kg delivered subcutaneously (s.c.) or intramuscularly (i.m).

The amount administered will depend on variables such as the type andextent of disease or indication to be treated, the overall health of thepatient, the in vivo potency of the antibody, the pharmaceuticalformulation, and the route of administration. The initial dosage can beincreased beyond the upper level in order to rapidly achieve the desiredblood-level or tissue level. Alternatively, the initial dosage can besmaller than the optimum, and the dosage may be progressively increasedduring the course of treatment. Human dosage can be optimized, e.g., ina conventional Phase I dose escalation study designed to run from, forexample, 0.5 mg/kg to 20 mg/kg. Dosing frequency can vary, depending onfactors such as route of administration, dosage amount and the diseasebeing treated. Exemplary dosing frequencies are more than once daily,about once per day, about twice a day, about three times a day, aboutfour times a day, about five times a day, about every other day, aboutevery third day, about once a week, about once every two weeks, aboutonce every month, about once every two months, about once every threemonths, about once every six months, or about once every year.Formulation of antibody-based drugs is within ordinary skill in the art.

In various embodiments, the antibodies of the invention may beadministered for a prolonged period. For example, the antibodies may beadministered for at least about 1 week, at least about 4 weeks, about 8weeks, or at least about 12 weeks. In some embodiments, the regimen isfor at least about 1 month, at least about 6 months, at least about 12months, at least about 1 year, at least about 2 years, at least about 3years, at least about 4 years, at least about 5 years, at least about 6years, at least about 7 years, at least about 8 years, at least about 9years, at least about 10 years, at least about 15 years, at least about20 years, at least about 30 years, at least about 40 years, or at leastabout 50 years.

For therapeutic use, an antibody may be combined with a pharmaceuticallyacceptable carrier. As used herein, “pharmaceutically acceptablecarrier” means buffers, carriers, and excipients suitable for use incontact with the tissues of human beings and animals without excessivetoxicity, irritation, allergic response, or other problem orcomplication, commensurate with a reasonable benefit/risk ratio. Thecarrier(s) should be “acceptable” in the sense of being compatible withthe other ingredients of the formulations and not deleterious to therecipient. Pharmaceutically acceptable carriers include buffers,solvents, dispersion media, coatings, isotonic and absorption delayingagents, and the like, that are compatible with pharmaceuticaladministration. The use of such media and agents for pharmaceuticallyactive substances is known in the art.

Pharmaceutical compositions containing antibodies, such as thosedisclosed herein, can be presented in a dosage unit form and can beprepared by any suitable method. A pharmaceutical composition should beformulated to be compatible with its intended route of administration.Examples of routes of administration are intravenous (IV), intradermal,inhalation, transdermal, topical, transmucosal, and rectaladministration. In an embodiment, the route of administration forantibodies of the invention is IV infusion. Useful formulations can beprepared by methods well known in the pharmaceutical art. For example,see Remington's Pharmaceutical Sciences, 18th ed. (Mack PublishingCompany, 1990).

In some embodiments, the pharmaceutical compositions are formulated as acomposition adapted for oral administration. Compositions for oraldelivery can be in the form of tablets, lozenges, aqueous or oilysuspensions, granules, powders, sprinkles, emulsions, capsules, syrups,or elixirs, for example. Orally administered compositions can compriseone or more agents, for example, sweetening agents such as fructose,aspartame or saccharin; flavoring agents such as peppermint, oil ofwintergreen, or cherry; coloring agents; and preserving agents, toprovide a pharmaceutically palatable preparation.

In some embodiments, the pharmaceutical compositions are formulated as acomposition adapted for parenteral administration. Dosage forms suitablefor parenteral administration (e.g. intravenous, subcutaneous,intramuscular, or intraperitoneal injection and infusion) include, forexample, solutions, suspensions, dispersions, emulsions, and the like.They may also be manufactured in the form of sterile solid compositions(e.g. lyophilized composition), which can be dissolved or suspended insterile injectable medium immediately before use. They may contain, forexample, suspending or dispersing agents.

In some embodiments, the compositions may additionally includepharmaceutically acceptable excipients or carriers. Exemplary excipientsinclude sodium citrate, dicalcium phosphate, etc., and/or a) fillers orextenders such as starches, lactose, sucrose, glucose, mannitol, silicicacid, microcrystalline cellulose, and Bakers Special Sugar, etc., b)binders such as, for example, carboxymethylcellulose, alginates,gelatin, polyvinylpyrrolidone, sucrose, acacia, polyvinyl alcohol,polyvinylpolypyrrolidone, methylcellulose, hydroxypropyl cellulose(HPC), and hydroxymethyl cellulose etc., c) humectants such as glycerol,etc., d) disintegrating agents such as agar-agar, calcium carbonate,potato or tapioca starch, alginic acid, certain silicates, sodiumcarbonate, cross-linked polymers such as crospovidone (cross-linkedpolyvinylpyrrolidone), croscarmellose sodium (cross-linked sodiumcarboxymethylcellulose), sodium starch glycolate, etc., e) solutionretarding agents such as paraffin, etc., f) absorption accelerators suchas quaternary ammonium compounds, etc., g) wetting agents such as, forexample, cetyl alcohol and glycerol monostearate, etc., h) absorbentssuch as kaolin and bentonite clay, etc., and i) lubricants such as talc,calcium stearate, magnesium stearate, solid polyethylene glycols, sodiumlauryl sulfate, glyceryl behenate, etc., and mixtures of suchexcipients. One of skill in the art will recognize that particularexcipients may have two or more functions.

Pharmaceutical formulations preferably are sterile. Sterilization can beaccomplished, for example, by filtration through sterile filtrationmembranes. Where the composition is lyophilized, filter sterilizationcan be conducted prior to or following lyophilization andreconstitution.

EXAMPLES Example 1. Construction and Characterization of HIV CrossMabAntibodies

FIGS. 13A-E and FIGS. 14A-E demonstrate that some iMab-based CrossMabshave greater potency and breadth than parental Abs. Except otherwisestated, all iMab-based bispecific antibodies were constructed using theMV1 variant. IC80, the antibody concentration that confers 80%neutralization of viral infectivity, is one method to evaluate antibodypotency against HIV. The lower the IC80 number (indicated in the y-axisof the graphs in term of antibody concentration (μg/ml)), the morepotent the antibody is at neutralizing a particular HIV strain orisolate. IC50, the antibody concentration that confers 50%neutralization of viral infectivity, is another method to evaluateantibody potency against HIV. The lower the IC50 number (indicated inthe y-axis of the graphs in term of antibody concentration (μg/ml)), themore potent the antibody is at neutralizing a particular HIV strain orisolate.

Various sets of antibodies were tested against a large panel of HIV-1pseudoviruses (118 different HIV viral isolates) representative of HIVenvelope diversity by geography, clade, tropism, and stage of infection.IC80 and IC50 were used to evaluate the strength of antiviral potencyand breadth. FIGS. 13E and 14E clearly demonstrate that, as compared tothe parental antibodies iMab and 10E8, the bispecific CrossMab of thetwo together (10E8/iMab) neutralizes almost all HIV viruses (each virusis indicated as a dot) more potently. The other antibody sets (used tomake 145/iMab, 117/iMab, 128/iMab and 151/iMab) sometimes enhance HIVpotency compared to their parental components and sometimes do not.

As shown in FIGS. 15A-E, the antibody iMab is also relatively potent incell-cell neutralizing assays. PGT145, 3BNC117, 10E8, PGT128 and PGT151are relatively potent at neutralizing cell-free viral infection, but arepoor in neutralizing viruses in cell-cell transmission assays. Creatingbispecific antibodies including PGT145, 3BNC117, 10E8, PGT128 and PGT151with iMab makes these chimeric antibodies active at neutralizing virusesin a cell-cell transmission assay. It can be seen that 10E8/iMab is themost potent antibody in these comparative studies. It is also found that10E8/iMab is most active in preventing cell-cell transmission in vitro.

As illustrated in FIG. 16, the improved potency of 10E8/iMab isstatistically significant. FIG. 3 shows that improved potency requirescovalent linkage of the antibody, i.e., the CrossMab format (sinceco-administration of two parental antibodies, iMab and 10E8, provides alower MPI than the fused and physically linked bispecific 10E8/iMabantibody). FIGS. 10, 11A-E, 12A-E, 13A-E, and 14A-E provide furtherevidence of the improved potency of iMab-derived CrossMab antibodiesover its parental antibodies.

In summary, it is found that, for the iMab-based CrossMabs (fused withPGT145, 3BNC117, PGT151, PGT128 and 10E8), 117/iMab improves breadth butnot potency; 145/iMab, 151/iMab and 128/iMab improve breadth andpotency; and 10E8/iMab markedly improves breadth and potency. In termsof epitope location/accessibility and potential models ofneutralization, 10E8/iMab appears to exhibit pre- and post-attachmentneutralization; 145/iMab, 151/iMab and 117/iMab appear to exhibitpre-attachment neutralization; and 117/iMab may show signs of stericrestriction and potentially reduced potency for some viruses. 10E8/iMabalso exhibits potent activity against HIV cell-to-cell transmission.

As also shown in FIGS. 13A-D and 14A-D, Pro 140-based CrossMabactivities are sometimes weaker than their parental antibodies andcorresponding iMab-based CrossMabs, as shown by the high concentrationsrequired to reach IC80 and IC50. Anchoring of these four mAbs to thehost cell receptor CCR5 via another host cell receptor-binding antibodycalled Pro 140 does not improve the antiviral potency or breadth (asmeasured by IC80 against a large panel of HIV isolates) compared totheir respective parental antibodies. These panels indicate thatPro140-based CrossMabs for these four antibodies are weaker than theircorresponding iMab-based CrossMabs (IC50 and IC80 comparisons ofPro140-based vs. iMab-based CrossMabs).

As shown in FIGS. 13E and 14E, 10E8/P140, a fifth Pro 140-basedCrossMab, is more potent than its parental antibodies and 10E8/iMabCrossMab. These panels illustrate a comparison of the potency (IC80 orIC50) of parental mAb Pro140 (right-most column of data points in FIGS.13E and 14E), bispecific CrossMab 10E8/P140 (second from right column ofdata points in FIGS. 13E and 14E), and parental mAb 10E8 (center columnof data points in FIGS. 13E and 14E) against a large panel of HIVisolates. These panels also illustrate a comparison of the potency (IC80or IC50) of parental mAb iMab (left-most column of data points in FIGS.13E and 14E), bispecific CrossMab 10E8/iMab (second from left column ofdata points in FIGS. 13E and 14E), and parental mAb 10E8 (center columnof data points in FIGS. 13E and 14E) against a large panel of HIVisolates. The second from left and second from right columns of datapoints in FIGS. 13E and 14E illustrate a comparison of the potency (IC80or IC50) of the bispecific CrossMabs 10E8/iMab and 10E8/P140 against alarge panel of HIV isolates.

Pro 140 is known to not have activity against X4 HIV viruses, as X4viruses use CXCR4 as a co-receptor for HIV-1 entry, and Pro 140 binds toCCR5. 10E8 alone has very weak activity against X4 viruses. However, thebispecific CrossMab 10E8/Pro 140 can neutralize all X4 viruses tested todate better than either of the parent antibodies. FIGS. 4A-J illustratethe effectiveness of 10E8, Pro 140, and 10E8/P140 bispecific CrossMabantibody in inhibition of various strains of HIV.

As shown in FIGS. 5A-G, 10E8/Pro140 CrossMab is a more potent inhibitorof various strains of HIV than the co-administration of the two parentalantibodies, demonstrating a synergistic, not merely additive,enhancement of potency with this particular bispecific antibody.

As shown in FIGS. 6A-D, a CrossMab of 10E8 fused to a non-membrane boundantibody (X19) does not provide enhanced potency, as can be seen whencompared to membrane bound 10E8/P140. Thus, the potency of the 10E8/P140CrossMab appears to require anchoring of 10E8 to the cell membrane.However, membrane binding alone does not afford the enhanced potency ofthese CrossMabs. FIGS. 7A-H show that anchoring 10E8 on HER2 does notprovide substantial potency enhancement as compared to anchoring 10E8 onCCR5. Anchoring of 10E8 to a viral receptor specifically (in this caseCCR5 via Pro 140 or CD4 via iMab) provides enhanced antiviral activity.

Δ10E8 is a mutant version of the 10E8 mAb that has a one amino aciddeletion in the light chain FR3. Compared to 10E8, Δ10E8 has a muchweaker epitope binding activity, as illustrated in FIGS. 8A-C. However,once the Δ10E8 was anchored on a cell receptor (by combining Δ10E8 andiMab in a CrossMab antibody—iMab specifically binds cell receptor CD4),FIGS. 8D-E, show that its inhibition activity is improved. These datasuggest the contribution of specific cell receptor anchoring, i.e.,anchoring on a viral receptor or a viral co-receptor, in enhancing theactivity of this HIV antibody. Still, while Δ10E8/P140 CrossMab hasimproved antiviral activity over Δ10E8, it is still not as potent as10E8/P140 CrossMab. Δ10E8/P140 CrossMab is comparatively more effectivein neutralizing R5 viruses than it is in neutralizing X4 viruses.

4E10 is an anti-gp41 MPER mAb known to be less potent than the anti-gp41MPER mAb 10E8. Similar to the results for Δ10E8, FIGS. 20 and 21A-G showthat anchoring 4E10 on co-receptor CCR5 (via Pro 140 in a CrossMabantibody) enhanced antiviral activity of 4E10 significantly. Takentogether, this suggests that the anchoring of a number of anti-gp41 MPERAbs to either CCR5 or CD4 (via combining the MPER Abs with P140 or iMabin a CrossMab antibody bispecific) can greatly improve the potency andbreadth of the respective anti-gp41 MPER Ab.

Multiple parameters contribute to enhanced activity of certainbispecific CrossMabs against HIV, including parental Ab potency,affinity, and pre- and post-attachment neutralization abilities. Inparticular, the 10E8/Pro140 CrossMab represents an effective combinationin terms of overcoming energetic, spatial and temporal constraints,targeting sequential/interdependent steps in the entry process, epitopelocation/accessibility, binding affinity, pre- and post-attachmentneutralization, and binding geometry. As shown in FIG. 20, 4E10/Pro140has a greater binding affinity for MPER than Δ10E8/Pro140 and10E8/Pro140. FIGS. 9A-G show the inhibition potency of 10E8/Pro140,Δ10E8/Pro140 and 4E10/Pro140, and their parental antibodies 10E8, Δ10E8and 4E10 against various strains of HIV. FIGS. 10, 13A-E, 14A-E, 15A-E,16, and 17A-B provide additional evidence of the greater potency ofCrossMab antibodies as compared to their parental antibodiesindividually and the parental antibodies in combination.

The enhanced antiviral coverage of 10E8/iMab and 10E8/Pro140 CrossMabsis illustrated in FIG. 10, which depict the potency and breadth ofseveral antibodies against HIV. The x-axis indicates the concentrationof a particular antibody, the y-axis indicates the percent of a largepanel of HIV viral isolates neutralized by a particular antibody at aspecific concentration, and each line indicates a different antibodyevaluated. The left-most lines along the x-axis and those that canclosely approach or reach 100% on the y-axis indicate a highly potentand broad antibody against HIV. 10E8/P140 CrossMab and 10E8/iMabCrossMab are among the most effective antibodies with respect to bothviral coverage and potency, and are significantly more effective thantheir parental antibodies.

FIGS. 18A-H and 19A-C show the potency of the CrossMab 10E8/515H7antibody as compared to its parental antibodies and previously discussedantibodies. The potency of a CrossMab antibody does not appear tocorrelate directly with the density of cell membrane protein targets, asthe density of CCR5 (the target of Pro140) is less than that of CD4 (thetarget of ibalizumab), yet the potency of 10E8/Pro140-derived CrossMabantibody is greater than that of 10E8/iMab-derived CrossMab antibody.

As shown in FIGS. 22A-B, the lack of single, sharp peaks in sizeexclusion chromatography indicates a type of instability indicative ofmultiple molecular species for 10E8 and 10E8-derived CrossMabantibodies. Table 1 recites various process and formulationmodifications used to resolve the 10E8 instability. However, asindicated by the “X” in the SEC or Size Exclusion Chromatography column,the modifications were unsuccessful in providing a single, sharp peak.

TABLE 1 Process and formulation screen to resolve 10E8 instabilityConditions SEC Purpose EDTA* X sequester metal ions, ↓ enzymaticactivity Acetic Acid* X ↓pH, stabilize protonated form of free thiols,↓reduction activity L Lysine* X competitive inhibitor against reductioncomponents CuSO4* X maintain reducing components in oxidized form,enzyme inhibitor 3-day harvesting X decreased cell death, ↓ enzymaticactivity SEC running X modification of analytical conditions buffercondition His formulation buffer X modification of analytical conditions

Pairing the 10E8 heavy chain with the light chain of 4E10 resolves theinstability issue, as shown in FIG. 23, producing a functional, thoughless potent, antibody. This result indicates that the instability of10E8 is due to the light chain. Various modifications of the 10E8 lightchain, shown in FIGS. 24B-C, such as removal of a C-terminal serine,engineering a lambda-variable region and kappa-constant region chimera,engineering an additional disulfide bond between the 10E8 heavy andlight chains, or genetically grafting kappa light chain regions ofnon-10E8 antibodies onto the 10E8 light chain do not fully resolve 10E8instability. As shown in FIGS. 25 and 26A-B, the instability is likelydue to a combination of the Complementarity Determining Regions (“CDRs”)and the framework regions (“FWs”) of 10E8. Using 10E8-HC/4E10-LC, each10E8 light chain CDR was grafted into 4E10LC individually or in concert,as shown in FIG. 25. Addition of 10E8 LC CDR2 and CDR3 are welltolerated, but addition of 10E8 LC CDR1 disrupts the single peak. Whenall 10E8 CDRs are grafted onto 4E10, the peak is broad. Grafting 10E8CDRs or frameworks onto its germline light chain λ results in a singlepeak, as shown in FIG. 26A, but effectiveness in MPER binding and HIVneutralization is decreased. Table 2 summarizes the 10E8 light chainvariants tested and the efficacy thereof

TABLE 2 Generated 10E8 LC variants MPER Neutral- ModificationsExpression binding SEC ization λLC′ → ΔS √ √ X √ λLC′ → κLC √ √ X √ LCCDR grafting (κLC Ab1) √ ↓ ? LC CDR grafting (κLC Ab2) √ ↓ ? H-L S—Sbond X X 10E8-H/4E10-L √ ↓ √ ↓ 10E8-H/4E 10-L CDR1 (10E8) X X 10E8-H/4E10-L CDR2 (10E8) √ ↓ √ ↓ 10E8-H/4E 10-L CDR3 (10E8) √ ↓ √ ↓ 10E8-H/4E10-L CDR123 (10E8) √ ↓ ? ↓

Variant H6L10 of 10E8 antibody was found to be active, non-autoreactive,and stable by size exclusion chromatography, as shown in FIG. 27.H6L10/Pro 140 and its parental antibodies were found to have comparablepharmacokinetics profiles in mice, as shown in FIG. 28. However, asshown in FIG. 29, the H6L10 variant of 10E8 (referred to as10E8_(v 1.0)) combined with P140 in a bispecific antibody issubstantially less potent than 10E8/P140 when tested against a largepanel of HIV strains. The H6L10 variant of 10E8 (referred to as10E8_(v 1.0)) combined with iMab in a bispecific antibody retains thesame relative amount of potency as compared to 10E8/iMab when testedagainst a large panel of HIV strains, but 10E8_(v 1.0)/iMab possessesthe same instability as 10E8/iMab as determined by size exclusionchromatography and indicated by an X in Table 3. In an embodiment, theH6L10 variant may further include a S74W mutation.

Table 3 below lists exemplary variants, their activities, size exclusionchromatography results, and pharmacokinetics (“PK”) results (see alsoFIG. 46).

TABLE 3 Exemplary variants that are stable while retaining anti-HIVactivity Construct Activity SEC PK 10E8/P140 +++++ X X 10E8_(v1.0/)P140++ √ √ 10E8_(v1.1/)P140 ++++ √ √ 10E8_(v2.0/)P140 +++ X ND10E8_(v3.0/)P140 +++++ √ X 10E8/iMab +++ X X 10E8_(v1.0/)iMab +++ X X10E8_(v1.1/)iMab +++ X X 10E8_(v2.0/)iMab ++++ √ √ 10E8_(v3.0/)iMab ++++√ X

As indicated above, 10E8_(V1.0) is a somatic variant of 10E8 known asH6L10. As a mAb, H6L10 has a single peak by SEC but reduced activitycompared to 10E8. H6L10/Pro140 CrossMab has single SEC peak and goodmouse PK, but reduced anti-HIV activity. H6L10/iMab CrossMab has doubleSEC peaks and poor mouse PK, but its activity against HIV is roughly thesame as 10E8/iMab. 10E8_(V1.1) includes a single point mutation inH6L10. When paired with Pro140 in a CrossMab bispecific, this constructhas single SEC peak and good mouse PK. Its activity against HIV isimproved as compared to 10E8_(V1.0)/Pro140, but still slightly less thanthat of 10E8/Pro140. When paired with iMab in a CrossMab bispecific,this construct has double SEC peaks and poor mouse PK, and its activityagainst HIV is still roughly the same as 10E8/iMab and 10E8_(V1.0)/iMab.10E8_(V2.0) is a chimeric antibody variant of 10E8 in which the FW1,CDR1 and part of FW2 are from 10E8_(V1.0) and in which the remainingpart of FW2, CDR2, FW3, CDR3 and FW4 are from 10E8. When paired withPro140 in a CrossMab bispecific, this construct has double SEC peaks andhas reduced activity against HIV as compared to 10E8/Pro140. When pairedwith iMab in a CrossMab bispecific, this construct has a single SECpeak, good PK, and activity against HIV that is improved over 10E8/iMab.10E8_(V3.0) is a somatic variant of 10E8 known as H11L1. H11L1/Pro140CrossMab has a single SEC peak and better anti-HIV activity than anyother 10E8/Pro140 construct (including the original one identified), buthas poor mouse PK due to autoreactivity. H11L1/iMab CrossMab has asingle SEC peak and anti-HIV activity that is better than the original10E8/iMab identified and roughly equivalent activity to that observedfor 10E8V2.0/iMab, but has poor mouse PK due to autoreactivity.

The variant of 10E8 that produced a single SEC peak in the context of aparticular CrossMab bispecific was different when paired with Pro140 oriMab. It appears that the stability of the 10E8 arm of these CrossMabbispecific antibodies is context dependent and will vary depending ofwhat antibody it is paired with. Thus, one variant (10E8_(V1.1)) wasidentified that was stable by SEC and with good mouse PK and goodanti-HIV activity when paired with Pro140. Another variant (10E8_(V2.0))was also identified that was stable by SEC with good mouse PK and withbetter anti-HIV activity than the originally identified 10E8/iMab.

Table 4 below describes the autoreactivity of tested variants, where“ANA” refers to anti-nuclear activity and “ACA” refers toanti-cardiolipin activity.

TABLE 4 Autoreactivity assessment in vitro Antibodies *Hep-2 (50 μg/mL)ANA ACA Staining score Negative control − − Low positive control + +High positive control ++++ ++++ iMab − − ′ Pro140 − − ′ 10E8_(v1.0) − −0 10E8_(v1.1) − − ′ 10E8_(v2.0) − − ′ 10E8_(v3.0) −/+ +/++ 0.510E8_(v1.0/)P140 − − ′ 10E8_(v2.0/)iMab − − ′ 10E8_(v1.1/)P140 − − ′10E8_(v3.0) ^(/)iMab − − ′ 10E8_(v3.0/)P140 − −/+ ′

FIG. 30 depicts the pharmacokinetics of 10E8 and CrossMab antibodiesderived from several 10E8 variants and iMab or P140 in a mouse model. Asshown in FIGS. 31A-D and 32A-B, 10E8_(v1.1/P)140 and 10E8_(v2.0)/iMabimprove anti-HIV activity and stability, and have good stability whenstored in PBS at 4° C. FIG. 33 depicts a native mass spectroscopyanalysis of 10E8_(v2.0)/iMab (N297A). FIGS. 34A-C compare the activityof 10E8_(v1.1/P)140 and 10E8_(v2.0)/iMab on a HIV Clade C panel, andcompares their IC50 and IC80 efficacy. FIGS. 35 and 36 compare thepotency of 10E8_(v1.1/P)140, 10E8_(v2.0)/iMab, and various monoclonalantibodies against HIV.

Example 2. Development of HIV CrossMab Antibodies with ImprovedSolubility, Stability, and/or Potency

Experiments were conducted to develop 10E8/iMab CrossMab antibodies withimproved solubility, stability, and activity.

Initially, a number of hydrophobic residues were identified on thesurface of the 10E8.2/iMab antibody (also referred to as the10E8_(v2.0)/iMab antibody) which may negatively affect solubility andstability of the bispecific antibody. The hydrophobic residues arepresented in Table 5 below (with reference to the Kabat numberingsystem):

TABLE 5 Location Residue L15 Leu L40 Pro L45 Ile L112 Pro H72 Leu H75Ile H77 Phe H89 Leu H98 Tyr H100a Phe H100b Trp H100e Tyr H100f ProH100i Pro H108 Leu H170 Leu

The hydrophobic residues were mutated either alone or in combination toyield 10E8.2/iMab variants that were assessed for their functionalactivity against HIV and in vivo pharmacokinetic profiles (see FIGS.37A-B). The amino acid sequences of the various 10E8.2/iMab variants areprovided elsewhere herein.

Specifically, in vitro neutralization assays were performed to test theactivity of the 10E8.2/iMab variants against HIV. Pseudoviruses wereprepared as previously described in Sun et al. 2014. JAIDS. 66, 473-483.Virus neutralization was assessed with a single cycle assay using TZM-blcells and HIV-1 pseudoviruses as described previously (Seaman et al.2010. J. Virol. 84, 1439-1452). As shown in FIG. 37A, some of the10E8.2/iMab variants (e.g., 10E8.2.1/iMab, 10E8.2.2/iMab, and10E8.2.3/iMab antibodies) retained functional activity in the in vitroHIV-1 neutralization assay as compared to the parental 10E8.2/iMabantibody.

For in vivo pharmacokinetics analysis, BALB/c mice were divided intogroups of three, and mice in each group were administeredintraperitoneally with 100 μg of the indicated antibody. Blood was drawnfrom all animals at Days 1, 2, 4, 7 and 10 post antibody administration,and serum was isolated and analyzed for levels of antibody in individualmice. CoStar 96-Well EIA/RIA plates were coated with 100 ng per well ofgoat anti-human IgG Fc-γ fragment overnight at 4° C. Plates were washedthree times with PBS+Tween and blocked with PBS containing 5% milk and0.5% BSA for 2 hours at room temperature. Mouse serum from the treatedanimals, and purified antibody in PBS for the standard curves, wereadded to the wells in 1:2 serial dilutions in PBS containing 2% milk and0.2% BSA and incubated for 2 hours. After washing, peroxidase-conjugatedgoat anti-human IgG was incubated for 1 hour at room temperature.Samples were detected by TMB Liquid Substrate System andspectrophotometric readings were performed at 450 nm. As shown in FIG.37B, some of the 10E8.2/iMab variants (e.g., 10E8.2.1/iMab,10E8.2.2/iMab, and 10E8.2.3/iMab antibodies) exhibited similarpharmacokinetic profiles as the parental 10E8.2/iMab antibody.

Additionally, the precipitation profiles of the variants were evaluatedunder thermal stress-inducing conditions. Specifically, 10E8.2/iMabvariants were expressed in 293 cells, purified using a Protein A column,exchanged into a solution of PBS (pH 7.4), and concentrated to >30 mg/mLusing a membrane with a nominal molecular weight limit of 50 kDa.Samples were then incubated at 50° C. and assessed visually forprecipitation at the indicated time points. Results from the thermalstress analysis are shown in FIG. 37C. These results indicate that the10E8.2.3/iMab variant retained the best combination of favorableantiviral activity by in vitro neutralization, favorable in vivopharmacokinetics, and increased solubility (decreased precipitation)under thermal stress inducing conditions.

Based on the favorable properties of 10E8.2.3/iMab in the precipitationassay, additional hydrophilic variants and combinations were created offof this bispecific antibody backbone variant. Each of these new variantswas evaluated for its aggregation potential by size exclusionchromatography (SEC) after incubation in thermal stress-inducingconditions (see FIGS. 38A-B). In particular, SEC was used to assess thephysicochemical homogeneity of the bispecific antibody variants and toresolve monomers from non-monomeric species. Results indicated that thevariants 10E8.4/iMab and 10E8.2.10/iMab exhibited the least aggregation,as indicated by a decreased peak size between 7 mL and 11 mL in FIG.38B. The 10E8.4/iMab variant was advanced into additional solubility andstability studies. This variant comprises a combination of 6 hydrophobicto hydrophilic residue mutations as compared to the parental 10E8.2/iMabantibody. FIGS. 45A-B provide a sequence alignment of the parental10E8.2/iMab antibody and the 10E8.4/iMab variant.

Based on the favorable functional and pharmacokinetic characteristicsand the reduction in precipitation and aggregation characteristics ofthe 10E8.4/iMab antibody, additional studies were performed to evaluatethe solubility, turbidity, thermostability and forced degradationcharacteristics of the 10E8.4/iMab antibody in comparison to the10E8.2/iMab antibody.

For example, the solubility of the 10E8.4/iMab antibody at 4° C. wasdetermined. In one set of experiments, the 10E8.2/iMab and 10E8.4/iMabantibodies were each buffer exchanged into target buffers and wereconcentrated via ultracentrifugation at 3000-5000 g, 4° C. Proteinconcentrations at different time points were determined by absorbance at280 nm using a NanoDrop 2000 spectrophotometer. All measurements wererepeated twice with 2.5 μL sample each time and an average was taken,and then protein concentration was plotted versus time. The maximumprotein concentration achieved was determined as the solubility of theprotein. As shown in FIGS. 39A-B, at concentrations above 50 mg/mL, the10E8.4/iMab antibody showed consistently higher protein concentrationsand solubility as compared to the 10E8.2/iMab antibody in buffers 1(acetate buffer, pH 4.5) and 2 (histidine buffer, pH 5.5).

The turbidity characteristics of the 10E8.4/iMab antibody were alsoanalyzed. In the analysis, the 10E8.2/iMab and 10E8.4/iMab antibodieswere each buffer exchanged into target buffers and were concentrated viaultracentrifugation at 3000-5000 g, 4° C. Absorbance at 280 nm and 350nm were measured over time using a NanoDrop 2000 spectrophotometer. Allmeasurements were repeated twice with 2.5 μL sample each time and anaverage was taken, and protein concentration (A280) was plotted versusturbidity (A350) for similar timepoints during the ultracentrifugationprocess. As shown in FIG. 40, the turbidity of both 10E8.2/iMab and10E8.4/iMab antibodies increased with protein concentration over time.In particular, the 10E8.2/iMab antibody showed higher turbidity than10E8.4/iMab at the same protein concentrations over 100 mg/mL in bothbuffer conditions tested.

Additionally, the thermostability profile of the 10E8.4/iMab antibodywas compared against the parental 10E8.2/iMab antibody usingdifferential scanning calorimetry (DSC). DSC is a thermoanalyticaltechnique in which the difference in the amount of heat required toincrease the temperature of a sample and reference is measured as afunction of temperature. Each peak in the thermogram corresponds to aheat effect associated with a specific process, such as crystallizationor melting, and is an indication of the stability of a molecule astemperature is increased. To determine the thermostability of the10E8.2/iMab and 10E8.4/iMab antibodies, each bispecific antibody wasbuffer exchanged into identical buffer compositions usingultra-filtration centrifugal devices under the condition of 4° C. and3000-5000 g. The protein concentrations were then adjusted to ˜10 mg/mLand aseptically filtered with 0.22-μm filter. Samples were then dilutedto 1 mg/mL with reference buffers. Reference buffers (400 μL) were addedinto the odd-numbered wells of a 96-well plate and 400 μL of sampleswere added into the even-numbered wells of the same plate. The plateswere scanned from 20° C. to 90° C. with a rate of 200° C./hr. Analysisof thermograms was performed with MicroCal VP-Capillary DSC

Automated data analysis software. As shown in FIG. 41, both 10E8.2/iMaband 10E8.4/iMab antibodies exhibited similar thermostability whenevaluated by DSC.

FIG. 42 shows the results from a turbidity analysis after forceddegradation of the 10E8.2/iMab and 10E8.4/iMab antibodies. The10E8.2/iMab and 10E8.4/iMab antibodies were each buffer exchanged intoidentical buffer compositions using ultra-filtration centrifugal devicesunder the condition of 4° C. and 3000-5000 g. The protein concentrationswere then adjusted to −10 mg/mL and aseptically filtered with 0.22-μmfilter. Samples were then incubated at 50° C. to induce forceddegradation, and both pre-centrifugation and post-centrifugation samplesof 10E8.2/iMab and 10E8.4/iMab were measured for development ofturbidity by absorbance at 350 nm at 0 days, 3 days and 6 days afterincubation began. Results indicate that the 10E8.2/iMab antibodyexhibited overall higher turbidity than 10E8.4/iMab at all time pointsstudied.

Molecule purity after forced degradation of the 10E8.2/iMab and10E8.4/iMab antibodies was also assessed. The 10E8.2/iMab and10E8.4/iMab antibodies were each buffer exchanged into two buffercompositions using ultra-filtration centrifugal devices under thecondition of 4° C. and 3000-5000 g. The protein concentrations were thenadjusted to ˜10 mg/mL and aseptically filtered with 0.22-μm filter.Samples were then incubated at 50° C. to induce forced degradation, andthe percentage of molecule purity by SDS-Gel Capillary Electrophoresisin non-reducing conditions was determined at 0 days, 3 days and 6 daysafter incubation began. To conduct SDS-Gel Capillary Electrophoresis, adenaturing solution was prepared by mixing sample buffer, 10% SDS and100 mM N-Ethylmaleimide at 20:1:0.7 volume ratio. Two microliters ofsamples and 7 μL denaturing solution were mixed well, incubated at 70°C. for 10 mins and spun down. 35 μL H₂O was added to the sample, then 42μL of the mixture was transferred into 96-well plate and centrifuged at4000 rpm for 20 mins to remove air bubbles. After the plate was loaded,samples were sipped, stained, separated and detected in the Microchipwhich was filled with destaining-gel, fluorescent dye and marker. Thedata was then analysed with LabChip GX Reviewer to determine thepercentage of intact bispecific antibody molecule versus smallerantibody fragments in each sample. As indicated in Table 6 below, the10E8.4/iMab antibody showed better intact molecule purity than the10E8.2/iMab antibody in both buffers at all timepoints studied:

TABLE 6 Purity (%) Formulation Timepoint 10E8.2/iMab 10E8.4/iMab Buffer#1 T0 71.6 87.6 Buffer #1 3 D 77.3 87.2 Buffer #1 6 D 76.2 87.0 Buffer#4 T0 74.9 89.2 Buffer #4 3 D 81.9 92.6 Buffer #4 6 D 89.4 94.6

An aggregation analysis was also performed. Specifically, the10E8.2/iMab and 10E8.4/iMab antibodies were each buffer exchanged intotwo buffer compositions using ultra-filtration centrifugal devices underthe condition of 4° C. and 3000-5000 g. The protein concentrations werethen adjusted to ˜10 mg/mL and aseptically filtered with 0.22-μm filter.Samples were then incubated at 50° C. to induce forced degradation, andthe high molecular weight (HMW) fraction in each protein sample wasdetermined by SE-HPLC (size exclusion chromatography) as a measurementof aggregation. Size exclusion chromatography was performed using anAgilent 1260 Infinity system and a TSKGel G3000SWXL column (300×7.8 mm,5 μm). The mobile phase was 50 mM PB, 300 mM NaCl, pH 7.0±0.2 and theflow rate was set as 1.0 mL/min. Samples were centrifuged (˜10000 rpmfor 2 min at 4° C.), injected and detected at 280 nm to determine thepercentage of HMW in the samples. As shown in Table 7 below, althoughthe 10E8.4/iMab antibody had a larger HMW population at TO, it showed aless rapid change in HMW percentage over time during incubation at theforced degradation-inducing conditions as compared to 10E8.2/iMab.

TABLE 7 HMW peak (%) Formulation Timepoint 10E8.2/iMab 10E8.4/iMabBuffer #1 T0 7.7 13.8 Buffer #1 3 D 39.2 27.2 Buffer #1 6 D 49.7 35.8Buffer #4 T0 5.9 11.0 Buffer #4 3 D 37.1 25.0 Buffer #4 6 D 44.9 31.3

Additionally, as shown in FIG. 43, the functional activities of the10E8.2/iMab and 10E8.4/iMab antibodies were compared in vitro. Inparticular, virus neutralization was assessed with a single cycle assayusing TZM-bl cells and 118 HIV-1 tier-2 HIV-1 Env pseudovirusesrepresenting diverse clades and origins as described previously (Seamanet al. 2010. J. Virol. 84, 1439-1452). Results indicate that, inaddition to its improvement in solubility, decrease in turbidity andimprovement in biophysical properties under thermal stress inducingconditions, the 10E8.4/iMab antibody also exhibited an approximately2.5-fold enhancement in neutralization activity against a large panel ofHIV-1 Env pseudotyped viruses as compared to 10E8.2/iMab.

The functional activities of the 10E8.2/iMab and 10E8.4/iMab antibodieswere also compared in vivo. Immunodeficient NSG mice (NOD.Cg-Prkdc^(scid) Il2rg^(tm1Wjl)/SzJ) were reconstituted with humanhematopoietic stem cells and infected with Tier-2 clade B HIV-1_(JR-CSF)four weeks prior to the initiation of antibody treatment. Mice were thentreated weekly with modified variants of 10E8.2/iMab or 10E8.4/iMab thatallowed for their evaluation in humanized mice. As shown in FIG. 44, amaximum mean viral load reduction of ˜1.7 log was observed in micetreated with 10E8.2/iMab, and a maximum mean viral load reduction of˜2.4 log was observed in mice treated with 10E8.4/iMab.”

The terms and expressions employed herein are used as terms andexpressions of description and not of limitation, and there is nointention, in the use of such terms and expressions, of excluding anyequivalents of the features shown and described or portions thereof. Inaddition, having described certain embodiments of the invention, it willbe apparent to those of ordinary skill in the art that other embodimentsincorporating the concepts disclosed herein may be used withoutdeparting from the spirit and scope of the invention. Accordingly, thedescribed embodiments are to be considered in all respects as onlyillustrative and not restrictive.

1. A bispecific antibody in a CrossMab format capable of neutralizingHIV, wherein the antibody comprises a light chain and heavy chainportion of a first antibody 10E8, or a variant thereof, that binds to aHIV envelope protein, and a light chain and heavy chain portion of asecond antibody ibalizumab, or a variant thereof, that binds to a cellmembrane receptor protein or a cell membrane co-receptor protein,wherein the light chain portion of the first antibody 10E8 comprises anamino acid sequence having at least 97% identity with SEQ ID NO: 33,wherein SEQ ID NO: 33 incorporates 1-4 mutations and the heavy chainportion of the first antibody 10E8 comprises an amino acid sequencehaving at least 97% identity with SEQ ID NO: 34, wherein SEQ ID NO: 34incorporates 4-12 mutations; and the light chain portion of the secondantibody ibalizumab comprises an amino acid sequence having at least 97%identity with SEQ ID NO: 1, and the heavy chain portion of the secondantibody ibalizumab comprises an amino acid sequence having at least 97%with SEQ ID NO: 2; and wherein any amino acid alterations relative toSEQ ID NOS: 1, 2, 33, and 34 are within the variable regions.
 2. Thebispecific antibody of claim 1, wherein the mutations to SEQ ID NO: 33are amino acid positions selected from L15, P40, I45, and/or P112. 3.The bispecific antibody of claim 1, wherein the mutations to SEQ ID NO:33 are amino acid positions selected from P40 and I45.
 4. The bispecificantibody of claim 1, wherein the mutations to SEQ ID NO: 33 are P4OT andI45K.
 5. The bispecific antibody of claim 1, wherein the mutations toSEQ ID NO: 34 are amino acid positions selected from L72, I75, F77, L89,Y98, F100a, W100b, Y100e, P100f, P100g, L108, and/or L170.
 6. Thebispecific antibody of claim 1, wherein the mutations to SEQ ID NO: 34are amino acid positions selected from L72, I75, F77, and/or L108. 7.The bispecific antibody of claim 1, wherein the mutations to SEQ ID NO:34 are L72K, I75K, F77T, and/or L108K.
 8. The bispecific antibody ofclaim 1, wherein the antibody comprises a light chain portion of anibalizumab antibody comprising the amino acid sequence of SEQ ID NO:
 19. The bispecific antibody of claim 1, wherein the antibody comprises aheavy chain portion of an ibalizumab antibody comprising the amino acidsequence of SEQ ID NO:
 2. 10. A pharmaceutical composition comprisingthe bispecific antibody of claim 1, and a pharmaceutically acceptablecarrier.
 11. The pharmaceutical composition of claim 10, wherein thecomposition is formulated for oral, intranasal, pulmonary, intradermal,transdermal, subcutaneous, intramuscular, intraperitoneal, orintravenous delivery.